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Genetic variation at the 19q13.3 KLK locus is linked with prostate cancer susceptibility in men. The non-synonymous KLK3 single nucleotide polymorphism (SNP), rs17632542 (c.536 T > C; Ile163Thr-substitution in PSA) is associated with reduced prostate cancer risk, however, the functional relevance is unknown. Here, we identify that the SNP variant-induced change in PSA biochemical activity mediates prostate cancer pathogenesis. The 'Thr' PSA variant leads to small subcutaneous tumours, supporting reduced prostate cancer risk. However, 'Thr' PSA also displays higher metastatic potential with pronounced osteolytic activity in an experimental metastasis in-vivo model. Biochemical characterisation of this PSA variant demonstrates markedly reduced proteolytic activity that correlates with differences in in-vivo tumour burden. The SNP is associated with increased risk for aggressive disease and prostate cancer-specific mortality in three independent cohorts, highlighting its critical function in mediating metastasis. Carriers of this SNP allele have reduced serum total PSA and a higher free/total PSA ratio that could contribute to late biopsy decisions and delay in diagnosis. Our results provide a molecular explanation for the prominent 19q13.3 KLK locus, rs17632542 SNP, association with a spectrum of prostate cancer clinical outcomes., (© 2024. The Author(s).)

Background: The kallikrein-kinin system (KKS) is a complex biochemical pathway that plays a crucial role in regulating several physiological processes, including inflammation, coagulation, and blood pressure. Dysregulation of the KKS has been associated with several pathological conditions such as hereditary angioedema (HAE), hypertension, and stroke. Developing an accurate quantitative model of the KKS may provide a better understanding of its role in health and disease and facilitate the rapid and targeted development of effective therapies for KKS-related disorders., Objectives: Here, we present a novel, detailed mechanistic model of the plasma KKS, elucidating the processes of Factor XII (FXII) activation, the kallikrein feedback loop, cleavage of high molecular weight kininogen leading to bradykinin (BK) production, and the impact of inhibitors., Methods: The model incorporates both surface and solution-phase reactions of all proteins in the KKS, describing how binding site concentration affects the rate of surface reactions. The model was calibrated and validated using a variety of published and in-house experimental datasets, which encompass a range of dextran sulphate (DXS) concentrations to initiate contact activation and various KKS inhibitors to block bradykinin production., Results: Our mathematical model showed that a trace amount of activated FXII is required for subsequent FXII activation. The model also reveals a bell-shaped curve relationship between the activation of the KKS and the number of DXS surface binding sites. Simulations of BK generation in healthy and HAE plasma demonstrated the impact of C1 esterase inhibitor (C1inh) deficiency via increased peak BK levels and accelerated formation in HAE plasma. The efficacy of KKS inhibitors, such as CSL312, ecallantide, and C1inh, was also evaluated, with CSL312 showing the most potent inhibition of BK generation., Conclusions: The present model represents a valuable framework for studying the intricate interactions within the plasma KKS and provides a better understanding of the mechanism of action of various KKS-targeted therapies., Competing Interests: I have read the journal’s policy, and the authors of this manuscript have the following competing interests: AR, HL, MP, CP, and IM are employees of CSL Limited. AR, HL, SD, MP, CP, and IM hold shares in CSL Limited., (Copyright: © 2024 Rezvani-Sharif et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Vascular endothelial injury is closely related to the progression of various cardio-cerebrovascular diseases. Whether Human Urinary Kallidinogenase (HUK) has a protective effect on endothelial injury remains unclear. This study established an in vivo model of rat common carotid artery intima injury and an in vitro model of human umbilical vein endothelial cell (HUVECs) injury induced by hydrogen peroxide (H 2 O 2 ). To explore the protective effect and mechanism of HUK on endothelial injury. In vivo, HUK can reduce the hyperplasia and lumen stenosis of rat common carotid artery after intimal injury, and promote the fluorescence expression of vWF in the common carotid artery. HUK also activated the Nrf2/HO-1 signaling pathway in rat common carotid artery tissue to reduce endothelial damage. In vitro, HUK can inhibit the H 2 O 2 -induced decline in HUVECs activity, improve the migration ability of HUVECs induced by H 2 O 2 , inhibit the apoptosis and necrosis of HUVECs and the generation of ROS, and regulate the expression of VEGFA, ET-1 and eNOS proteins related to endothelial function in cells. The Nrf2/HO-1 signaling pathway is activated, and the HO-1 specific inhibitor zinc porphyrin (ZnPP) can partially reverse the protective effect of HUK on H 2 O 2 -induced HUVECs injury in terms of cell migration, necrosis and oxidative stress. The Nrf2/HO-1 signaling pathway plays an important role in the regulation of migration, necrosis and oxidative stress of HUVECs cells. HUK has a protective effect on vascular endothelial injury. HUK can inhibit oxidative stress and apoptotic necrosis by activating Nrf2/HO-1 signaling pathway., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Alzheimer's disease (AD) and related dementias constitute an important global health challenge. Detailed understanding of the multiple molecular mechanisms underlying their pathogenesis constitutes a clue for the management of the disease. Kallikrein-related peptidases (KLKs), a lead family of serine proteases, have emerged as potential biomarkers and therapeutic targets in the context of AD and associated cognitive decline. Hence, KLKs were proposed to display multifaceted impacts influencing various aspects of neurodegeneration, including amyloid-beta aggregation, tau pathology, neuroinflammation, and synaptic dysfunction. We propose here a comprehensive survey to summarize recent findings, providing an overview of the main kallikreins implicated in AD pathophysiology namely KLK8, KLK6 and KLK7. We explore the interplay between KLKs and key AD molecular pathways, shedding light on their significance as potential biomarkers for early disease detection. We also discuss their pertinence as therapeutic targets for disease-modifying interventions to develop innovative therapeutic strategies aimed at halting or ameliorating the progression of AD and associated dementias., (Copyright © 2024 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Matrix metalloproteinases (MMPs) are involved in the breakdown of lung extracellular matrix and the consequent release of Mycobacterium tuberculosis into the airways. Recent studies indicate that kallikrein-related peptidase 12 (KLK12) regulate MMP-1 and MMP-9, suggesting that targeting the KLK12 gene could be a promising tuberculosis (TB) treatment. To maximise therapeutic potential, this strategy of silencing KLK12 needs to be delivered to the pathogenic cell population while preserving the immunoprotective and tissue homeostatic functions of other lung macrophages. Our research found that KLK12 is highly expressed in M2 macrophages, leading us to design mannose-based bovine serum albumin nanoparticles (MBNPs) for delivering siRNA to silence KLK12 in these cells. The results of in vitro experiments showed that MBNPs could accurately enter M2 macrophages and sustainably release KLK12-siRNA with the help of mannose and mannose receptor targeting. The results of the in vivo experiments showed that MBNPs could reach the lungs within 1 h after intraperitoneal injection and peaked at 6 h. MBNPs increased collagen fibre content in the lungs by decreasing the levels of KLK12/MMPs thereby limiting the progression of TB. Importantly, MBNPs provided greater alleviation of pulmonary TB symptoms and reduced bacterial load in both TB and drug-resistant TB models. These findings provide an alternative and effective option for the treatment of TB, especially when drug resistance occurs. STATEMENT OF SIGNIFICANCE: RNA interference using small interfering RNA (siRNA) can target various genes and has potential for treating diseases such as tuberculosis (TB). However, siRNAs are unstable in the blood and within cells. This study presents bovine serum albumin nanoparticles encapsulating KLK12-siRNA (BNPs) synthesized via desolvation. A mannose layer was added (MBNPs) to target mannose receptors on M2 macrophages, facilitating endocytosis. The low pH-responsive MBNPs enhance lysosomal escape for siRNA delivery, downregulating the KLK12 pathway. Tests confirmed that MBNPs effectively inhibited Mycobacterium bovis proliferation, reduced granulomas, and decreased inflammation in a mouse model. This research aims to reduce antibiotic use, shorten treatment duration, and provide a novel TB treatment option., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

The widespread presence of microplastics in the environment has raised significant concerns regarding their potential impact on human and animal health. Among various microplastic types, polyethylene microplastics (PE-MPs) are particularly prevalent due to the extensive use in packaging and consumer products. Exploring the uncharted therapeutic potentials of naringin, this study delves into its mitigating effects on disruptions in kallikrein-3 levels, steroidal-thyroidal hormone balance, and antioxidant defense triggered by PE-MPs exposure, paving the way for novel interventions in environmental toxin-induced endocrine and oxidative stress disorders. Male Wistar rats (n = 24) were randomly grouped into four: Control, PE-MPs (1.5 mg/kg), PE-MPs + NAR (1.5 mg/kg PE-MPs + 100 mg/kg NAR), and NAR (100 mg/kg). Hormonal and antioxidant parameters were assessed after 28 days of exposure. PE-MPs exposure caused a significant increase(p < 0.005) in the level of kallikrein-3 (KLK-3) while it significantly reduces the levels of testosterone (TST), luteinizing hormone, thyroid stimulating hormone (TSH) and Free-triiodothyronine (fT3) and Total cholesterol (TChol) concentration. PE-MPs exposure also disrupted significantly (p < 0.005) antioxidant profile by down-regulating the activities of glutathione-S-transferase, catalase (CAT), superoxide dismutase (SOD) and reducing levels of glutathione (GSH) and ascorbic acid (AA) while concentration of malondialdehyde (MDA) levels were increased relative to control. However, the mitigating potentials of naringin on disruptions in hormonal and antioxidant profiles caused by PE-MPs exposure were demonstrated, as NAR normalized KLK-3, steroid, and thyroid hormone levels, cholesterol concentration, and enhanced antioxidant defense. This suggests that NAR is a promising protective agent against endocrine and oxidative damage induced by environmental contaminants such as microplastics., (© 2024. The Author(s).)

Introduction: Human kallikrein-related peptidases (KLKs) represent a subgroup of 15 serine endopeptidases involved in various physiological processes and pathologies, including cancer., Areas Covered: This review aims to provide a comprehensive overview of the KLK family, highlighting their genomic structure, expression profiles and substrate specificity. We explore the role of KLKs in tumorigenesis, emphasizing their potential as biomarkers and therapeutic targets in cancer treatment. The dysregulated activity of KLKs has been linked to various malignancies, making them promising candidates for cancer diagnostics and therapy., Expert Opinion: : Recent advancements in understanding the mechanistic pathways of KLK-related tumorigenesis offer new prospects for developing targeted cancer treatments. Expert opinion suggests that while significant progress has been made, further research is necessary to fully exploit KLKs' potential in clinical applications.

The human kallikrein-related peptidase (KLK) family which consists of 15 members is associated with prostate cancer and other cancers. It has been reported that overexpression of KLK4 in prostate cancer correlates with bone metastasis or advanced stage. Hypoxia occurs in the early stages of prostate cancer due to the accumulation of acidic metabolites or reactive oxygen species (ROS). In our study, KLK4 gene expression in hypoxic conditions in PC-3 and LNCaP cells which are treated with TGF-β was evaluated with mRNA, protein, and promoter activity levels. A chemical hypoxia model was created and confirmed at mRNA and protein level. No statistically significant cytotoxic effect of CoCl 2 and TGF-β was observed in PC-3 and LNCaP cells with the MTT test. Four different truncated KLK4 gene promoter constructs were cloned in pmetLuc expression vector and basal activities of all promoter fragments were analyzed. The activities of P1 (-447/ + 657), P2 (-103/ + 657), and P3 (-267/ + 657) promoter fragments increased in hypoxic conditions except P4 (+555/ + 657), which does not contain the SMAD and HRE region. KLK4 mRNA levels in both PC-3 and LNCaP cells increased in the hypoxia and hypoxia/TGF groups compared to the non-treated groups. The stimulating effect of TGF-β is correlated with the increase in SMAD2/3 mRNA levels. KLK4 expression is up-regulated by TGF-β, especially under hypoxic conditions, and its interaction with the SMAD pathway is determined with different inhibitor experiments. HIF-1α and SMAD transcription factors bind to the KLK4 promoter showing the direct interaction of HIF-1α (-80/-52) and SMAD (+163/+194) regions with EMSA., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Hypertension is one of the main risk factors for cardiovascular disease and causes widespread morbidity and mortality worldwide. The aim of this work was to screen the fruit with high angiotensin I-converting enzyme (ACE) inhibitory activity and kallikrein (KLK) promotion activity by three different extraction methods from 22 kinds of fruits. Results showed that the aqueous extracts of fresh kiwifruit significantly inhibited ACE activity (47.71%), whereas the KLK activity was also inhibited (4.56%). This indicated that the substances inhibiting ACE activity existed in kiwifruit might be small molecular substances such as polyphenols. The nonpolar substance existed in the ethanol extracts of grape inhibited ACE activity significantly. The enzymatic hydrolysates of red grape significantly promoted KLK activity, whereas its ethanol extracts significantly inhibited KLK activity. This results suggested that the components that lower blood pressure and raise blood pressure are generally presented in the same fruit, the former are mostly water-soluble substances, while the latter are generally alcohol-soluble substances. If certain or individual components can be isolated from edible fruits, they may significantly affect blood pressure in humans., Competing Interests: Declarations Ethics Approval Not applicable. Consent to Participate The present paper has been approved by all named authors. Consent for publication The present paper, which is original, has not been published before and is not currently being considered for publication elsewhere. Competing Interests The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Coronaviruses rely on host proteases to activate the viral spike protein, which facilitates fusion with the host cell membrane and the release of viral genomic RNAs into the host cell cytoplasm. The distribution of specific host proteases in the host determines the host, tissue, and cellular tropism of these viruses. Here, we identified the kallikrein (KLK) family member KLK5 as a major host protease secreted by human airway cells and exploited by multiple human betacoronaviruses. KLK5 cleaved both the priming (S1/S2) and activation (S2') sites of spike proteins from various human betacoronaviruses in vitro. In contrast, KLK12 and KLK13 displayed preferences for either the S2' or S1/S2 site, respectively. Whereas KLK12 and KLK13 worked in concert to activate SARS-CoV-2 and MERS-CoV spike proteins, KLK5 by itself efficiently activated spike proteins from several human betacoronaviruses, including SARS-CoV-2. Infection of differentiated human bronchial epithelial cells (HBECs) with human betacoronaviruses induced an increase in KLK5 that promoted virus replication. Furthermore, ursolic acid and other related plant-derived triterpenoids that inhibit KLK5 effectively suppressed the replication of SARS-CoV, MERS-CoV, and SARS-CoV-2 in HBECs and mitigated lung inflammation in mice infected with MERS-CoV or SARS-CoV-2. We propose that KLK5 is a pancoronavirus host factor and a promising therapeutic target for current and future coronavirus-induced diseases.

The kallikrein-related peptidase KLK2 has restricted expression in the prostate luminal epithelium, and its protein target is unknown. The present work reports the hydrolytic activities of KLK2 on libraries of fluorescence resonance energy-transfer peptides from which the sequence SYRIF was the most susceptible substrate for KLK2. The sequence SYRIF is present at the extracellular N -terminal segment ( 58 SYRIF 63 Q) of IL-10R2. KLK2 was fully active at pH 8.0-8.2, found only in prostate inflammatory conditions, and strongly activated by sodium citrate and glycosaminoglycans, the quantities and structures controlled by prostate cells. Bone-marrow-derived macrophages (BMDM) have IL-10R2 expressed on the cell surface, which is significantly reduced after KLK2 treatment, as determined by flow cytometry (FACS analysis). The IL-10 inhibition of the inflammatory response to LPS/IFN-γ in BMDM cells due to decreased nitric oxide, TNF-α, and IL-12 p40 levels is significantly reduced upon treatment of these cells with KLK2. Similar experiments with KLK3 did not show these effects. These observations indicate that KLK2 proteolytic activity plays a role in prostate inflammation and makes KLK2 a promising target for prostatitis treatment.

Backgrounds: KLKs have been proved to be key regulators of the tumor microenvironment. In this study, we explored the potential of Kallikrein-related peptidases (KLKs) as clinical diagnostic and prognostic markers in patients with kidney renal clear cell carcinoma (KIRC) as well as their relationship with common immuno-inhibitor and immune cell infiltration in the tumor microenvironment to provide new targets and novel ideas for KIRC therapy., Methods: Oncomine, Gene Expression Profiling Interactive Analysis (GEPIA), UCSC Xena, Genotype-Tissue Expression (GTEx), Kaplan-Meier plotter, cBioPortal, STRING, GeneMANIA, and TISIDB were used to analyze the differential expression, prognostic value, gene changes, molecular interaction, and immune infiltration of KLKs in patients with KIRC., Results: From the gene expression level, it can be determined that KLK1, KLK6, and KLK7 are differentially expressed in KIRC and normal tissues. From the perspective of clinical prognosis, KLK1, KLK13, and KLK14 are highly correlated with the clinical prognosis of KIRC. The expression of KLKs is regulated by various immunosuppressive agents, with KDR, PVRL2, and VTCN1 being the most significant. The expression of KLKs is significantly correlated with the infiltration of various immune cells, of which Eosinophils and Neutrophils are the most significant., Conclusions: KLK1, KLK6, KLK7, KLK13, and KLK14 have potential as diagnostic and prognostic biomarkers, among which KLK1 is the most significant. This study may provide detailed immune information and promising targets for KIRC immunotherapy to assist in designing new immunotherapies., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Wang et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Human tissue kallikrein-related peptidase 7 (KLK7) is a serine protease implicated in the physiology of skin desquamation, and its uncontrolled activity can lead to chronic diseases such as psoriasis, atopic dermatitis, and Netherton syndrome. For this reason, kallikrein 7 has been identified as a potential therapeutic target. This work aimed to evaluate Pluronic (PL) hydrogels as topical carriers of four specific scFv-Fc antibodies to inhibit KLK7. The hydrogels comprised PL F127 (30% w/v) alone and a binary F127/P123 (28-2% w/v) system. Each formulation was loaded with 1 μg/mL of each antibody and characterized by physicochemical and pharmaceutical techniques, considering antibody-micelle interactions and hydrogel behavior as smart delivery systems. Results showed that the antibodies were successfully loaded into the PL-based systems, and the sol-gel transition temperature was shifted to high values after the P123 addition. The antibodies released from the gels preserved their rheological properties ( G ' > G '', 35- to 41-fold) and inhibitory activity against KLK7, even after 24 h. This work presented potential agents targeting KLK7 that may provide strategies for treating skin abnormalities.

Background: Kallikrein-related peptidase 7 (KLK7) is a chymotrypsin-like serine protease which is essential for the desquamation of corneocytes and thus plays a pivotal role in maintaining skin homeostasis. In cancer, KLK7 overexpression was suggested to represent a route for metastasis through cleavage of cell junction and extracellular matrix proteins of cancer cells., Methods: To comprehensively determine KLK7 protein expression in normal and neoplastic tissues, a tissue microarray containing 13,447 samples from 147 different tumor types and subtypes as well as 608 samples of 76 different normal tissue types was analyzed by immunohistochemistry., Results: KLK7 positivity was found in 64 of 147 tumor categories, including 17 tumor categories with at least one strongly positive case. The highest rate of KLK7 positivity was found in squamous cell carcinomas from various sites of origin (positive in 18.1%-63.8%), ovarian and endometrium cancers (4.8%-56.2%), salivary gland tumors (4.8%-13.7%), bilio-pancreatic adenocarcinomas (20.0%-40.4%), and adenocarcinomas of the upper gastrointestinal tract (3.3%-12.5%). KLK7 positivity was linked to nodal metastasis (p = 0.0005), blood vessel infiltration (p = 0.0037), and lymph vessel infiltration (p < 0.0001) in colorectal adenocarcinoma, nodal metastasis in hepatocellular carcinoma (p = 0.0382), advanced pathological tumor stage in papillary thyroid cancer (p = 0.0132), and low grade of malignancy in a cohort of 719 squamous cell carcinomas from 11 different sites of origin (p < 0.0001)., Conclusions: These data provide a comprehensive overview on KLK7 expression in normal and neoplastic human tissues. The prognostic relevance of KLK7 expression and the possible role of KLK7 as a drug target need to be further investigated., (© 2024. The Author(s).)

Background: Lympho-epithelial Kazal-type-related inhibitor (LEKTI) is a serine protease inhibitor consisting of multiple domains. A loss of function mutation is described in Netherton patients that show severe symptoms of atopic lesions and itch., Objectives: LEKTI domain 6 (LD6) has shown strong serine protease-inhibitory action in in vitro assays and thus it was tested in vitro and in vivo for potential anti-inflammatory action in models of atopic skin disease., Methods: Human skin equivalents were treated with LD6 and an inflammatory reaction was challenged by kallikrein-related endopeptidase 5 (KLK5). Furthermore, LD6 was tested on dorsal root ganglia cells stimulated with KLK5, SLIGRL and histamine by calcium imaging. The effect of topically administered LD6 (0.4-0.8%) in lipoderm was compared to a topical formulation of betamethasone-diproprionate (0.1%) in a therapeutic setting on atopic dermatitis-like lesions in NC/Nga mice sensitized to house dust mite antigen. Endpoints were clinical scoring of the mice as well as determination of scratching behaviour., Results: KLK5 induced an upregulation of CXCL-8, CCL20 and IL-6 in skin equivalents. This upregulation was reduced by pre-incubation with LD6. KLK5 as well as histamine induced calcium influx in a population of neurons. LD6 significantly reduced the calcium response to both stimuli. When administered onto lesional skin of NC/Nga mice, both LD6 and betamethasone-dipropionate significantly reduced the inflammatory reaction. The effect on itch behaviour was less pronounced., Conclusion: Topical administration of LD6 might be a new therapeutic option for treatment of lesional atopic skin., Competing Interests: Declaration of Competing Interest The authors have no conflict of interest to declare, (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Advancing healthcare for elderly men requires a deeper understanding of testicular aging processes. In this study, we conducted transcriptomic profiling of 43,323 testicular single cells from young and old mice, shedding light on 1032 telocytes-an underexplored testicular cell type in previous research. Our study unveiled 916 age-related differentially expressed genes (age-DEGs), with telocytes emerging as the cell type harboring the highest count of age-DEGs. Of particular interest, four genes (Klk1b21, Klk1b22, Klk1b24, Klk1b27) from the Kallikrein family, specifically expressed in Leydig cells, displayed down-regulation in aged testes. Moreover, cell-type-level splicing analyses unveiled 1838 age-related alternative splicing (AS) events. While we confirmed the presence of more age-DEGs in somatic cells compared to germ cells, unexpectedly, more age-related AS events were identified in germ cells. Further experimental validation highlighted 4930555F03Rik, a non-coding RNA gene exhibiting significant age-related AS changes. Our study represents the first age-related single-cell transcriptomic investigation of testicular telocytes and Kallikrein genes in Leydig cells, as well as the first delineation of cell-type-level AS dynamics during testicular aging in mice., (© 2024. The Author(s).)

Liver cancer is one of the most pivotal global health problems, leading hepatocellular carcinoma (HCC) with a significant increase in cases worldwide. The role of non-coding-RNA in cancer proliferation and carcinogenesis has attracted much attention in the last decade; however, microRNAs (miRNAs), as non-coding RNA, are considered master mediators in various cancer progressions. Yet the role of miR-141 as a modulator for specific cellular processes in liver cancer cell proliferation is still unclear. This study identified the role of miR-141 and its potential functions in liver carcinogenesis. The level of miR-141 in HepG2 and HuH7 cells was assessed using quantitative real-time PCR (qRT-PCR) and compared with its expression in normal hepatocytes. A new miR-141 construct has been performed in a CMV promoter vector tagged with GFP. Using microarray analysis, we identified the potentially regulated genes by miR-141 in transfected HepG2 cells. The protein profile of the kallikrein-related peptidase 10 (KLK10) and tumor necrosis factor TNFSF-15 was investigated in HepG2 cells transfected with either an inhibitor, antagonist miR-141, or miR-141 overexpression vector using immunoblotting and flow cytometry assay. Finally, ELISA assay has been used to monitor the produced inflammatory cytokines from transfected HepG2 cells. Our findings showed that the expression of miR-141 significantly increased in HepG2 and HuH7 cells compared to the normal hepatocytes. Transfection of HepG2 cells with an inhibitor, antagonist miR-141, showed a significant reduction of HepG2 cell viability, unlike the transfection of miR-141 overexpression vector. The microarray data of HepG2 cells overexpressed miR-141 provided a hundred downregulated genes, including KLK10 and TNFSF-15. Furthermore, the expression profile of KLK10 and TNFSF-15 markedly depleted in HepG2 cells transfected with miR-141 overexpression accompanied by a decreasing level of interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF-α), indicating the role of miR-141 in HepG2 cell proliferation and programmed cell death. Interestingly, the experimental rats with liver cancer induced by Diethylnitrosamine injection further confirmed the upregulation of miR-141 level, IL-10, and TNF-α and the disturbance in KLK10 and TNFSF-15 gene expression compared with their expression in normal rats. The in-silico online tools, IntaRNA and miRWalk were used to confirm the direct interaction and potential binding sites between miR-141 and identified genes. Thus, the seeding regions of potential targeted sequences was cloned upstream of luciferase reporter gene in pGL3 control vector. Interestingly, the luciferase activities of constructed vectors were significantly decreased in HepG2 cells pre-transfected with miR-141 overexpression vector, while increasing in cells pre-transfected with miR-141 specific inhibitor. In summary, these data suggest the crucial role of miR-141 in liver cancer development via targeting KLK10 and TNFSF-15 and provide miR-141 as an attractive candidate in liver cancer treatment and protection., (© 2024. The Author(s).)

Squamous cell carcinoma is the most common primary tumor in the head and neck epithelium and is the second most common primary tumor type in the lung. Although morphologically indistinguishable from each other with hematoxylin and eosin stain on histology, the tumors have different protein expression profiles. Using 24 formalin-fixed paraffin embedded squamous cell carcinomas of the lung and 24 squamous cell carcinomas in the head and neck, protein expression for cytokeratin 5/6, kallikrein 7, and elafin was evaluated by immunohistochemistry. All three proteins were found to evidence higher expression in head and neck squamous cell carcinoma as compared with that of squamous cell carcinoma of the lung. The differences in expression may help clinical differentiation between primary tumors of the lung from metastatic tumors to the lung from the oral/laryngeal cavities.

Prolylcarboxypeptidase (PRCP, PCP, Lysosomal Pro-X-carboxypeptidase, Angiotensinase C) controls angiotensin- and kinin-induced cell signaling. Elevation of PRCP appears to be activated in chronic inflammatory diseases [cardiovascular disease (CVD), diabetes] in proportion to severity. Vascular endothelial cell senescence and mitochondrial dysfunction have consistently been shown in models of CVD in aging. Cellular senescence, a driver of age-related dysfunction, can differentially alter the expression of lysosomal enzymes due to lysosomal membrane permeability. There is a lack of data demonstrating the effect of age-related dysfunction on the expression and function of PRCP. To explore the changes in PRCP, the PRCP-dependent prekallikrein (PK) pathway was characterized in early- and late-passage human pulmonary artery endothelial cells (HPAECs). Detailed kinetic analysis of cells treated with high molecular weight kininogen (HK), a precursor of bradykinin (BK), and PK revealed a mechanism by which senescent HPAECs activate the generation of kallikrein upon the assembly of the HK-PK complex on HPAECs in parallel with an upregulation of PRCP and endothelial nitric oxide (NO) synthase (eNOS) and NO formation. The NO production and expression of both PRCP and eNOS increased in early-passage HPAECs and decreased in late-passage HPAECs. Low activity of PRCP in late-passage HPAECs was associated with rapid decreased telomerase reverse transcriptase mRNA levels. We also found that, with an increase in the passage number of HPAECs, reduced PRCP altered the respiration rate. These results indicated that aging dysregulates PRCP protein expression, and further studies will shed light into the complexity of the PRCP-dependent signaling pathway in aging.

Heart failure (HF) is a pervasive clinical challenge characterized by compromised cardiac function and reduced quality of life. The kinin-kallikrein system (KSS), a multifaceted peptide cascade, has garnered substantial attention due to its potential role in HF. Through activation of B1 and/or B2 receptors and downstream signaling, kinins modulate various physiological processes, including inflammation, coagulation, pain, blood pressure control, and vascular permeability. Notably, aberrations in KKS components have been linked to HF risk. The elevation of vasodilatory bradykinin (BK) due to kallikrein activity reduces preload and afterload, while concurrently fostering sodium reabsorption inhibition. However, kallikrein's conversion of prorenin to renin leads to angiotensinsII upregulation, resulting in vasoconstriction and fluid retention, alongside increased immune cell activity that fuels inflammation and cardiac remodeling. Importantly, prolonged KKS activation resulting from volume overload and tissue stretch contributes to cardiac collagen loss. The conventional renin-angiotensin-aldosterone system (RAAS) inhibitors used in HF management may inadvertently intensify KKS activity, exacerbating collagen depletion and cardiac remodeling. It is crucial to balance the KKS's role in acute cardiac damage, which may temporarily enhance function and metabolic parameters against its detrimental long-term effects. Thus, KKS blockade emerges as a promising strategy to impede HF progression. By attenuating the link between immune system function and tissue damage, KKS inhibition can potentially reduce cardiac remodeling and alleviate HF symptoms. However, the nuanced roles of BK in various acute conditions necessitate further investigation into the sustained benefits of kallikrein inhibitors in patients with chronic HF., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Tumor endothelial cells (TECs), which are thought to be structurally and functionally different from normal endothelial cells (NECs), are increasingly attracting attention as a therapeutic target in hypervascular malignancies. Although colorectal liver metastasis (CRLM) tumors are hypovascular, inhibitors of angiogenesis are a key drug in multidisciplinary therapy, and TECs might be involved in the development and progression of cancer. Here, we analyzed the function of TEC in the CRLM tumor microenvironment. We used a murine colon cancer cell line (CT26) and isolated TECs from CRLM tumors. TECs showed higher proliferation and migration than NECs. Coinjection of CT26 and TECs yielded rapid tumor formation in vivo. Immunofluorescence analysis showed that coinjection of CT26 and TECs increased vessel formation and Ki-67 + cells. Transcriptome analysis identified kallikrein-related peptide 10 (KLK10) as a candidate target. Coinjection of CT26 and TECs after KLK10 downregulation with siRNA suppressed tumor formation in vivo. TEC secretion of KLK10 decreased after KLK10 downregulation, and conditioned medium after KLK10 knockdown in TECs suppressed CT26 proliferative activity. Double immunofluorescence staining of KLK10 and CD31 in CRLM tissues revealed a significant correlation between poor prognosis and positive KLK10 expression in TECs and tumor cells. On multivariate analysis, KLK10 expression was an independent prognostic factor in disease-free survival. In conclusion, KLK10 derived from TECs accelerates colon cancer cell proliferation and hematogenous liver metastasis formation. KLK10 in TECs might offer a promising therapeutic target in CRLM., (© 2024 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.)

Kirsten rat sarcoma virus (KRAS) gene mutation is common in colorectal cancer (CRC) and is often predictive of treatment failure and poor prognosis. To understand the mechanism, we compared the transcriptome of CRC patients with wild-type and mutant KRAS and found that KRAS mutation is associated with the overexpression of a secreted serine protease, kallikrein-related peptidase 10 (KLK10). Moreover, using in vitro and in vivo models, we found that KLK10 overexpression favors the rapid growth and liver metastasis of KRAS mutant CRC and can also impair the efficacy of KRAS inhibitors, leading to drug resistance and poor survival. Further functional assays revealed that the oncogenic role of KLK10 is mediated by protease-activated receptor 1 (PAR1). KLK10 cleaves and activates PAR1, which further activates 3-phosphoinositide-dependent kinase 1 (PDK1)-AKT oncogenic pathway. Notably, suppressing PAR1-PDK1-AKT cascade via KLK10 knockdown can effectively inhibit CRC progression and improve the sensitivity to KRAS inhibitor, providing a promising therapeutic strategy. Taken together, our study showed that KLK10 promotes the progression of KRAS mutant CRC via activating PAR1-PDK1-AKT signaling pathway. These findings expanded our knowledge of CRC development, especially in the setting of KRAS mutation, and also provided novel targets for clinical intervention., (© 2023 International Federation for Cell Biology.)

Nummular eczema, a chronic dermatitis characterized by coin-shaped lesions, was first documented in 1857. However, its pathophysiological characteristics are still not well known. To investigate differences in the regulation of the desquamation process in the stratum corneum of lesional and nonlesional skin of patients with nummular eczema and healthy control subjects, tape-stripped stratum corneum samples from patients with nummular eczema and healthy volunteers were analysed using immunofluorescence staining and western blot analysis. In the nummular eczema lesional skin, expression of desmoglein-1, desmocollin-1, and corneodesmosin exhibited a disorganized, dense or partially diffuse non-peripheral pattern with increased intensity, compared with the peripheral patterns observed in healthy or nonlesional skin, suggesting the impaired desquamation process in nummular eczema. Furthermore, although the expression of the desquamation-related serine proteases, kallikrein-related peptidase 7 and 5, was increased in nummular eczema lesional skin, the immunofluorescence staining of lympho-epithelial Kazal-type-related inhibitor-1, an endogenous inhibitor of various kallikrein-related peptidases, and its fragments were significantly increased in the nummular eczema lesional skin, suggesting its contribution to the inhibition of corneodesmosomal degradation. Therefore, the increased detection of corneodesmosomal proteins in nummular eczema lesions may be due to the increased amount of the fragments of lympho-epithelial Kazal-type-related inhibitor-1, which could contribute to delayed desquamation.

The identification of surfaceome proteins is a main goal in cancer research to design antibody-based therapeutic strategies. T cell engagers based on KLK2, a kallikrein specifically expressed in prostate cancer (PRAD), are currently in early clinical development. Using genomic information from different sources, we evaluated the immune microenvironment and genomic profile of prostate tumors with high expression of KLK2. KLK2 was specifically expressed in PRAD but it was not significant associated with Gleason score. Additionally, KLK2 expression did not associate with the presence of any immune cell population and T cell activating markers. A mild correlation between the high expression of KLK2 and the deletion of TMPRSS2 was identified. KLK2 expression associated with high levels of surface proteins linked with a detrimental response to immune checkpoint inhibitors (ICIs) including CHRNA2, FAM174B, OR51E2, TSPAN1, PTPRN2, and the non-surface protein TRPM4. However, no association of these genes with an outcome in PRAD was observed. Finally, the expression of these genes in PRAD did not associate with an outcome in PRAD and any immune populations. We describe the immunologic microenvironment on PRAD tumors with a high expression of KLK2, including a gene signature linked with an inert immune microenvironment, that predicts the response to ICIs in other tumor types. Strategies targeting KLK2 with T cell engagers or antibody-drug conjugates will define whether T cell mobilization or antigen release and stimulation of immune cell death are sufficient effects to induce clinical activity.

During early postnatal brain development, the formation of proper synaptic connections between neurons is crucial for the development of functional neural networks. Recent studies have established the involvement of protease-mediated modulations of extracellular components in both synapse formation and elimination. The secretory serine protease neuropsin (also known as kallikrein-8) cleaves a few transmembrane or extracellular matrix proteins in a neural activity-dependent manner and regulates neural plasticity. However, neuropsin-dependent proteolysis of extracellular components and the involvement of these components in mouse brain development are poorly understood. We have observed that during hippocampus development, expression of neuropsin and levels of full-length or cleaved fragments of the neuropsin substrate protein L1 cell adhesion molecule (L1CAM) positively correlate with synaptogenesis. Our subcellular fractionation studies show that the expression of neuropsin and its proteolytic activity on L1CAM are enriched at developing hippocampal synapses. Activation of neuropsin expression upregulates the transcription and cleavage of L1CAM. Furthermore, blocking of neuropsin activity, as well as knockdown of L1CAM expression, significantly downregulates in vitro hippocampal synaptogenesis. Taken together, these findings provide evidence for the involvement of neuropsin activity-dependent regulation of L1CAM expression and cleavage in hippocampal synaptogenesis., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Porcine placental extract (PPE) is commonly used in various health foods and cosmetics. PPE use in cosmetics predominantly consist of the water-soluble fraction derived from the entire placenta. In this report, we examined the effect of the hydrophobic constituents of the PPE, specifically the sphingolipid-enriched fraction designated as the sphingolipid-enriched porcine placental extract (SLPPE), on the expression of genes associated with skin function in cultured normal human epidermal keratinocytes. Using quantitative RT-PCR (qRT-PCR) analysis, we found that SLPPE concentrations ranging from 25 to 100 µg/mL upregulated the gene expression of key components associated with the cornified envelope structure (filaggrin (FLG), involucrin (IVL) and loricrin (LOR)), cornification enzymes (transglutaminase 1 (TGM1) and TGM5) and the desquamation enzymes (kallikrein 5 (KLK5) and KLK7). Additionally, KLK5p and FLG protein (FLGp) were detected in the culture supernatants of keratinocytes treated with SLPPE at these concentrations. These findings suggest that SLPPE is possible to promote the cornification and desquamation in epidermal keratinocytes, and it may offer potential benefits in cosmetics.

Endometriosis is a common benign gynecologic condition. Endometriosis lesions are associated with endometrial cell proliferation, migration, invasion, and neovascularization, while the specific molecular mechanisms are still elusive. Transcriptome sequencing has been used for the identification of diagnostic markers in endometriosis. Here, transcriptome profiling revealed that kallikrein-related peptidase 4 (KLK4) expression was up-regulated in ectopic endometrium (EC) tissues of patients with endometriosis. KLK4 mediates the degradation of extracellular matrix proteins, and its proteolytic activity activates many tumorigenic and metastatic pathways via tumor invasion and migration. Nevertheless, whether KLK4 serves as an important regulatory factor in endometriosis remains unclear. This study confirmed that KLK4 was highly expressed in ectopic endometrial stromal cells (EC-ESCs). KLK4 overexpression promoted proliferation and suppressed apoptosis of EC-ESCs, induced cell migration and invasion, and enhanced angiogenesis in vivo. Mechanistically, KLK4 overexpression mediated the protein cleavage of pro-brain-derived neurotrophic factor in EC-ESCs. Finally, brain-derived neurotrophic factor was a vital downstream substrate of KLK4 maintained the proliferation, metastasis, and pro-angiogenesis abilities and inhibited apoptosis of ESCs through a rescue study. Together, these findings demonstrate the promotive role of KLK4 in endometriosis development. In addition, the study provides a new insight that KLK4 might be a potential therapeutic target and prognostic marker for patients with endometriosis., (Copyright © 2024 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

The incidence and mortality rates of colorectal cancer (CRC) have significantly increased in recent years. It has been shown that early diagnosis of CRC improves the five-year survival of patients compared to late diagnosis, as patients with stage I disease have a five-year survival rate as high as 90 %. Through bioinformatics analysis, we identified Kallikrein 10 (KLK10), a member of the Kallikrein family, as a reliable predictor of CRC progression, particularly in patients with early-stage CRC. Furthermore, single-cell analysis revealed that KLK10 was highly expressed in tumor and partial immune cells. Analysis of the biological functions of KLK10 using the Kyoto encyclopedia of genes and genomes and gene ontology indicated that KLK10 plays a role in the proliferation and differentiation of cancer cells, along with the maintenance of tumor function and immune regulation, explicitly by T cells and macrophages. EdU cell proliferation staining, plate clone formation assay, and cell scratch assay demonstrated that KLK10 inhibition by siRNA affected the proliferation and migration of CRC cells. Cell cycle detection by flow cytometry demonstrated that KLK10 inhibition led to cell cycle arrest in the G1 phase. In addition, the proportion of M1 and M2 macrophages in 45 tumor specimens was analyzed by immunohistochemistry, the proportion of CD4 + T cells and CD8 + T cells in plasma was identified by flow cytometry, and their correlation with KLK10 was analyzed. The effects of KLK10 on T cells and macrophages were verified in independent cell experiments. The results revealed that KLK10 also activates CD4 + T cells, mediating M2-type macrophage polarization., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Investigation on a competitive endogenous RNA (ceRNA) network attracted lots of attention due its function in cancer regulation. Here, we probed into the possible molecular mechanism of circSSPO/microRNA-6820-5p (miR-6820-5p)/kallikrein-related peptidase 8 (KLK8)/PKD1 network in the esophageal squamous cell carcinoma (ESCC). Following whole-transcriptome sequencing and differential analysis in collected ESCC tissue samples, circRNA-miRNA-mRNA regulatory network affecting ESCC was investigated. After interaction measurement among circSSPO/miR-6820-5p/KLK8/PKD1, their regulatory roles in ESCC cell functions in vitro and xenograft tumor growth and lung metastasis in vivo were analyzed. The bioinformatics prediction and sequencing results screened that circSSPO, miR-6820-5p, KLK8, and PKD1 were associated with ESCC development. In ESCC, miR-6820-5p was expressed at very low levels, while circSSPO, KLK8, and PKD1 were highly expressed. In vitro cell experiments further proved that circSSPO competitively inhibited miR-6820-5p to induce ESCC cell malignant properties. Moreover, knockdown of KLK8 or PKD1 inhibited ESCC cell malignant properties. circSSPO also promoted the tumorigenic and metastasis of ESCC through the upregulation of KLK8 and PKD1 expression in vivo. We found that circSSPO was an oncogenic circRNA that was significantly abundant in ESCC tissues and circSSPO exhibited an oncogenic activity in ESCC by elevating expression of KLK8 and PKD1 through suppressing miR-6820-5p expression., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Introduction: Endothelial injury is a major characteristic of sepsis and contributes to sepsis-induced multiple-organ dysfunction. In this study, we investigated the role of miR-107-3p in sepsis-induced endothelial injury., Methods: Human umbilical vein endothelial cells (HUVECs) were exposed to 20 μg/mL of lipopolysaccharide (LPS) for 6-48 h. The levels of miR-107-3p and kallikrein-related peptidase 5 (KLK5) were examined. HUVECs were treated with LPS for 12 h and subsequently transfected with miR-107-3p inhibitor, KLK5 siRNA, or cotransfected with KLK5 siRNA and miR-107-3p inhibitor/negative control inhibitor. Cell survival, apoptosis, invasion, cell permeability, inflammatory response, and the Toll-like receptor 4/nuclear factor κB signaling were evaluated. In addition, the relationship between miR-107-3p and KLK5 expression was predicted and verified., Results: LPS significantly elevated miR-107-3p levels, which peaked at 12 h. Conversely, the KLK5 level was lower in the LPS group than in the control group and was lowest at 12 h. MiR-107-3p knockdown significantly attenuated reductions in cell survival and invasion, apoptosis promotion, hyperpermeability and inflammation induction, and activation of the NF-κB signaling caused by LPS. KLK5 knockdown had the opposite effect. Additionally, KLK5 was demonstrated as a target of miR-107-3p. MiR-107-3p knockdown partially reversed the effects of KLK5 depletion in LPS-activated HUVECs., Conclusions: Our findings indicate that miR-107-3p knockdown may protect against sepsis-induced endothelial cell injury by targeting KLK5. This study identified a novel therapeutic target for sepsis-induced endothelial injury., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Axonal injury and demyelination occur in demyelinating diseases, such as multiple sclerosis, and the detachment of myelin from axons precedes its degradation. Paranodes are the areas at which each layer of the myelin sheath adheres tightly to axons. The destruction of nodal and paranodal structures during inflammation is an important pathophysiology of various neurological disorders. However, the underlying pathological changes in these structures remain unclear. Kallikrein 6 (KLK6), a serine protease produced by oligodendrocytes, is involved in demyelinating diseases. In the present study, we intraperitoneally injected mice with LPS for several days and examined changes in the localization of KLK6. Transient changes in the intracellular localization of KLK6 to paranodes in the spinal cord were observed during LPS-induced systemic inflammation. However, these changes were not detected in the upper part of brain white matter. LPS-induced changes were suppressed by minocycline, suggesting the involvement of microglia. Moreover, nodal lengths were elongated in LPS-treated wild-type mice, but not in LPS-treated KLK6-KO mice. These results demonstrate the potential involvement of KLK6 in the process of demyelination., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

We aimed to study mRNA levels and prognostic impact of all 15 human kallikrein-related peptidases (KLKs) and their targets, proteinase-activated receptors (PARs), in surgically treated prostate cancer (PCa). Seventy-nine patients with localized grade group 2-4 PCas represented aggressive cases, based on metastatic progression during median follow-up of 11 years. Eighty-six patients with similar baseline characteristics, but no metastasis during follow-up, were assigned as controls. Transcript counts were detected with nCounter technology. KLK12 protein expression was investigated with immunohistochemistry. The effects of KLK12 and KLK15 were studied in LNCaP cells using RNA interference. KLK3, -2, -4, -11, -15, -10 and -12 mRNA, in decreasing order, were expressed over limit of detection (LOD). The expression of KLK2, -3, -4 and -15 was decreased and KLK12 increased in aggressive cancers, compared to controls (P < .05). Low KLK2, -3 and -15 expression was associated with short metastasis-free survival (P < .05) in Kaplan-Meier analysis. PAR1 and -2 were expressed over LOD, and PAR1 expression was higher, and PAR2 lower, in aggressive cases than controls. Together, KLKs and PARs improved classification of metastatic and lethal disease over grade, pathological stage and prostate-specific antigen combined, in random forest analyses. Strong KLK12 immunohistochemical staining was associated with short metastasis-free and PCa-specific survival in Kaplan-Meier analysis (P < .05). Knock-down of KLK15 reduced colony formation of LNCaP cells grown on Matrigel basement membrane preparation. These results support the involvement of several KLKs in PCa progression, highlighting, that they may serve as prognostic PCa biomarkers., (© 2023 The Authors. International Journal of Cancer published by John Wiley & Sons Ltd on behalf of UICC.)

In order to study mechanistic/mammalian target of rapamycin's role in T cell differentiation, we generated mice in which Rheb is selectively deleted in T cells (T-Rheb-/- C57BL/6J background). During these studies, we noted that T-Rheb-/- mice were consistently heavier but had improved glucose tolerance and insulin sensitivity as well as a marked increase in beige fat. Microarray analysis of Rheb-/- T cells revealed a marked increase in expression of kallikrein 1-related peptidase b22 (Klk1b22). Overexpression of KLK1b22 in vitro enhanced insulin receptor signaling, and systemic overexpression of KLK1b22 in C57BL/6J mice also enhances glucose tolerance. Although KLK1B22 expression was markedly elevated in the T-Rheb-/- T cells, we never observed any expression in wild-type T cells. Interestingly, in querying the mouse Immunologic Genome Project, we found that Klk1b22 expression was also increased in wild-type 129S1/SVLMJ and C3HEJ mice. Indeed, both strains of mice demonstrate exceptionally improved glucose tolerance. This prompted us to employ CRISPR-mediated knockout of KLK1b22 in 129S1/SVLMJ mice, which in fact led to reduced glucose tolerance. Overall, our studies reveal (to our knowledge) a novel role for KLK1b22 in regulating systemic metabolism and demonstrate the ability of T cell-derived KLK1b22 to regulate systemic metabolism. Notably, however, further studies have revealed that this is a serendipitous finding unrelated to Rheb., (Copyright © 2023 The Authors.)

Objective: Patients with rheumatoid arthritis (RA) have shown increased levels of neutrophils generating kallikrein-kinin peptides in blood which are potent mediators of inflammation. This study investigated the association between the bioregulation of kinin-mediated inflammation with the clinical, quality of life, and imaging characteristics (e.g. ultrasonography) of different arthritides., Methods: Patients with osteoarthritis (OA, n = 29), gout (n = 10) and RA (n = 8) were recruited and screened for clinical symptoms, quality of life, and ultrasonographical assessment of arthritis. Blood neutrophils were assessed for the expression of bradykinin receptors (B1R and B2R), kininogens and kallikreins by immunocytochemistry with visualization by bright field microscopy. Levels of plasma biomarkers were measured by ELISA and cytometric bead array., Results: Quality of life (SF-36 domains and summary scores; including pain; and, HAQ) was similar across OA, gout and RA patients; with the exception of worse physical functioning scores between OA and gout patients. Synovial hypertrophy (on ultrasound) differed between groups (p = 0.001), and the dichotomised Power Doppler (PD) score of greater than or equal to 2 (PD-GE2) was marginally significant (p = 0.09). Plasma IL-8 were highest in patients with gout followed by RA and OA (both, P < 0.05). Patients with RA had higher plasma levels of sTNFR1, IL-1β, IL-12p70, TNF and IL-6, compared to OA and gout patients (all, P < 0.05). Patients with OA had higher expression of K1B and KLK1 on blood neutrophils followed by RA and gout patients (both, P < 0.05). Bodily pain correlated with B1R expression on blood neutrophils (r = 0.334, p = 0.05), and inversely with plasma levels of CRP (r = -0.55), sTNFR1 (r = -0.352) and IL-6 (r = -0.422), all P < 0.05. Expression of B1R on blood neutrophils also correlated with Knee PD (r = 0.403) and PD-GE2 (r = 0.480), both P < 0.05., Conclusions: Pain levels and quality of life were similar between patients with OA, RA and gout with knee arthritis. Plasma inflammatory biomarkers and B1R expression on blood neutrophils correlated with pain. Targeting B1R to modulate the kinin-kallikrein system may pose as a new therapeutic target in the treatment of arthritis., (© 2023. The Author(s).)

Kallikrein-related peptides (KLKs) form an evolutionally conserved subgroup of secreted serine proteases that consists of 15 members (KLK1-15). Previous studies have shown that KLKs regulate diverse biological processes, but the clinical significance of KLKs remains largely unclear in human breast cancers. We examined the expression profile of 15 KLK genes in breast carcinomas using microarray data. Next, we immunolocalized KLK12 in 140 breast carcinomas and evaluated its clinical significance. Subsequently, we examined the effects of KLK12 on proliferation and migration in breast cancer cell lines. From microarray analyses, it turned out that KLK12 was the most strongly associated with low-grade malignancy in breast carcinomas among the 15 KLK members. Immunohistochemical KLK12 status was positively associated with ER and PR status, while it was inversely associated with stage, pathological T factor, lymph node metastasis, and distant metastasis. Prognostic analyses demonstrated that KLK12 was a favorable prognostic factor for both disease-free and breast cancer-specific survival of the patients. Furthermore, the knockdown of KLK12 significantly increased cell proliferation activity and cell migration of breast cancer cells. These results suggest that KLK12 has antitumorigenic effects associated with proliferation and migration and immunohistochemical KLK12 status as a potent favorable prognostic factor in breast carcinoma patients.

Neuronal apoptosis has been well-recognized as a critical mediator in the pathogenesis of depressive disorders. Tissue kallikrein-related peptidase 8 (KLK8), a trypsin-like serine protease, has been implicated in the pathogenesis of several psychiatric disorders. The present study aimed to explore the potential function of KLK8 in hippocampal neuronal cell apoptosis associated with depressive disorders in rodent models of chronic unpredictable mild stress (CUMS)-induced depression. It was found that depression-like behavior in CUMS-induced mice was associated with hippocampal KLK8 upregulation. Transgenic overexpression of KLK8 exacerbated, whereas KLK8 deficiency attenuated CUMS-induced depression-like behaviors and hippocampal neuronal apoptosis. In HT22 murine hippocampal neuronal cells and primary hippocampal neurons, adenovirus-mediated overexpression of KLK8 (Ad-KLK8) was sufficient to induce neuron apoptosis. Mechanistically, it was identified that the neural cell adhesion molecule 1 (NCAM1) may associate with KLK8 in hippocampal neurons as KLK8 proteolytically cleaved the NCAM1 extracellular domain. Immunofluorescent staining exhibited decreased NCAM1 in hippocampal sections obtained from mice or rats exposed to CUMS. Transgenic overexpression of KLK8 exacerbated, whereas KLK8 deficiency largely prevented CUMS-induced loss of NCAM1 in the hippocampus. Both adenovirus-mediated overexpression of NCAM1 and NCAM1 mimetic peptide rescued KLK8-overexpressed neuron cells from apoptosis. Collectively, this study identified a new pro-apoptotic mechanism in the hippocampus during the pathogenesis of CUMS-induced depression via the upregulation of KLK8, and raised the possibility of KLK8 as a potential therapeutic target for depression., (© 2023. The Author(s).)

Background: Human serum albumin (HSA) is the most abundant plasma protein and is sensitive to glycation in vivo. The chronic hyperglycemic conditions in patients with diabetes mellitus (DM) induce a nonenzymatic Maillard reaction that denatures plasma proteins and forms advanced glycation end products (AGEs). HSA-AGE is a prevalent misfolded protein in patients with DM and is associated with factor XII activation and downstream proinflammatory kallikrein-kinin system activity without any associated procoagulant activity of the intrinsic pathway., Objectives: This study aimed to determine the relevance of HSA-AGE toward diabetic pathophysiology., Methods: The plasma obtained from patients with DM and euglycemic volunteers was probed for activation of FXII, prekallikrein (PK), and cleaved high-molecular-weight kininogen by immunoblotting. Constitutive plasma kallikrein activity was determined via chromogenic assay. Activation and kinetic modulation of FXII, PK, FXI, FIX, and FX via in vitro-generated HSA-AGE were explored using chromogenic assays, plasma-clotting assays, and an in vitro flow model using whole blood., Results: Plasma obtained from patients with DM contained increased plasma AGEs, activated FXIIa, and resultant cleaved cleaved high-molecular-weight kininogen. Elevated constitutive plasma kallikrein enzymatic activity was identified, which positively correlated with glycated hemoglobin levels, representing the first evidence of this phenomenon. HSA-AGE, generated in vitro, triggered FXIIa-dependent PK activation but limited the intrinsic coagulation pathway activation by inhibiting FXIa and FIXa-dependent FX activation in plasma., Conclusion: These data indicate a proinflammatory role of HSA-AGEs in the pathophysiology of DM via FXII and kallikrein-kinin system activation. A procoagulant effect of FXII activation was lost through the inhibition of FXIa and FIXa-dependent FX activation by HSA-AGEs., Competing Interests: Declaration of competing interests There are no competing interests to disclose., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Enamel is formed by the repetitive secretion of a tooth-specific extracellular matrix and its decomposition. Calcification of the enamel matrix via hydroxyapatite (HAP) maturation requires pH cycling to be tightly regulated through the neutralization of protons released during HAP synthesis. We found that Gpr115, which responds to changes in extracellular pH, plays an important role in enamel formation. Gpr115-deficient mice show partial enamel hypomineralization, suggesting that other pH-responsive molecules may be involved. In this study, we focused on the role of Gpr111/Adgrf2, a duplicate gene of Gpr115, in tooth development. Gpr111 was highly expressed in mature ameloblasts. Gpr111-KO mice showed enamel hypomineralization. Dysplasia of enamel rods and high carbon content seen in Gpr111-deficient mice suggested the presence of residual enamel matrices in enamel. Depletion of Gpr111 in dental epithelial cells induced the expression of ameloblast-specific protease, kallikrein-related peptidase 4 (Klk4), suggesting that Gpr111 may act as a suppressor of Klk4 expression. Moreover, reduction of extracellular pH to 6.8 suppressed the expression of Gpr111, while the converse increased Klk4 expression. Such induction of Klk4 was synergistically enhanced by Gpr111 knockdown, suggesting that proper enamel mineralization may be linked to the modulation of Klk4 expression by Gpr111. Furthermore, our in vitro suppression of Gpr111 and Gpr115 expression indicated that their suppressive effect on calcification was additive. These results suggest that both Gpr111 and Gpr115 respond to extracellular pH, contribute to the expression of proteolytic enzymes, and regulate the pH cycle, thereby playing important roles in enamel formation., (© 2023 Federation of American Societies for Experimental Biology.)

Skin surface pH has been identified as a key regulator of the epidermal homeostasis through its action on serine protease activity. These enzymes, like kallikreins (KLK), are responsible for the degradation of corneodesmosomes, the protein structures linking together corneocytes, and are regulated by Lympho-Epithelial Kazal-Type-related Inhibitor (LEKTI). KLK activity increases at pH levels higher than physiological. An increase in skin surface pH has been observed in patients suffering from skin diseases characterized by impaired barrier function, like atopic dermatitis. In this work, we introduce an agent-based model of the epidermis to study the impact of a change in skin surface pH on the structural and physiological properties of the epidermis, through the LEKTI-KLK mechanism. We demonstrate that a less acidic pH, compared to the slightly acidic pH observed in healthy skin, is sufficient to significantly affect the water loss at the surface and the amount of irritant permeating through the epidermis. This weakening of the skin barrier function eventually results in a more intense skin inflammation following exposure to an external irritant. This work provides additional evidence that skin surface pH and serine proteases can be therapeutic targets to improve skin barrier integrity., (© 2022 John Wiley & Sons Ltd.)

The renal kallikrein-kinin system (RKKS) has been related to blood pressure control and sodium and water balance. We have previously shown that female spontaneously hypertensive rats (SHR) have high urinary kallikrein activity (UKa) and lower blood pressure (BP) than males whereas ovariectomy stimulates UKa and diminishes BP. We also showed that high K + intake and prepuberal gonadectomy (Gx) diminish BP with a concomitant increase in UKa and plasma aldosterone levels. Since kallikrein co-localize in the same distal nephron segments of aldosterone effectors, we explored the effect of pharmacological blockage of aldosterone receptor, epithelial Na + (ENaC) and the rectifying outer medulla K + (ROMK) channels in different gonad contexts on the gene expression, renal tissue content and urine release of kallikrein. Klk1 gene expression was determined by real-time PCR and enzymatic activity of kallikrein by the amidolytic method. We found that the inhibition of the aldosterone receptor by spironolactone increases kallikrein renal tissue storage and decreases its urinary activity, especially in Gx rats. Moreover, ENaC blockade by benzamil increases the renal content of kallikrein without affecting synthesis or excretion, especially in females and Gx animals, while the inhibition of ROMK by glibenclamide increases the synthesis and renal content of kallikrein only in intact male animals. We concluded that RKKS regulation showed sexual dimorphism and seemed to be modulated by sex hormones throughout a process involving aldosterone and the aldosterone-sensitive ion channels.., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: Mendelian disorders of cornification (MeDOC) are a group of heterogeneous genodermatoses with different genetic bases. The pathogenesis of a substantial group of MeDOC remains to be elucidated., Objectives: To identify a new causative gene and the pathogenesis of a previously undescribed autosomal-dominant cornification disorder., Methods: Whole-exome sequencing was performed in three families with the novel cornification disorder to identify the disease-causing variants. As the variants were located around the signal peptide (SP) cleavage site of a kallikrein-related peptidase, SP cleavage, subcellular localization and extracellular secretion of the variants were evaluated in eukaryotic overexpression systems by Western blotting or immunocytochemistry. Then the trypsin-like and chymotrypsin-like proteolytic activity of the peptidase and degradation of its catalytic substrate were assayed using the patients' stratum corneum (SC) samples. The morphology of the lamellar bodies and corneodesmosomes (CDs) in the patients' SC was ultrastructurally examined. A mouse model harbouring the equivalent variant was constructed and evaluated histologically., Results: We identified two heterozygous variants affecting Gly50 in kallikrein-related peptidase (KLK)11 in a familial case and two sporadic cases with the new disorder, which is characterized by early-onset ichthyosiform erythroderma or erythrokeratoderma. KLK11 belongs to the family of kallikrein-related peptidases participating in skin desquamation by decomposing CDs, a process essential for shedding of the SC. In vitro experiments demonstrated that the variants perturbed the SP cleavage of KLK11, leading to subcellular mislocalization and impaired extracellular secretion of the KLK11 Gly50Glu variant. Both trypsin-like and chymotrypsin-like proteolytic activities were significantly decreased in the patients' SC samples. Reduced proteolysis of desmoglein 1 and delayed degeneration of CDs were detected in patients' SC, indicating delayed skin desquamation. Consistently, the patients showed a thickened, dense SC, indicating abnormal skin desquamation. Mice harbouring the homozygous c.131G>A (p.Gly44Glu) Klk11 variant, which is equivalent to KLK11 c.149G>A (p.Gly50Glu) in humans, exhibited hyperkeratosis and abnormal desquamation, partially recapitulating the phenotype., Conclusions: We provide evidence that variants at Gly50 affecting the SP cleavage of KLK11 cause a new autosomal-dominant cornification disorder with abnormal desquamation. Our findings highlight the essential role of KLKs in maintaining homeostasis of skin keratinization and desquamation., Competing Interests: Conflicts of interest: The authors declare they have no conflicts of interest., (© The Author(s) 2022. Published by Oxford University Press on behalf of British Association of Dermatologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

The vast majority of the human proteome is yet to be functionally characterized thus hindering ongoing investigations on potential drug resistance mechanisms and advanced treatment options. Chemical proteomics is a powerful solution for enzyme profiling and the development of next generation cancer therapeutics previously deemed undruggable by small molecules. Within this field, activity-based protein profiling (ABPP) is a specialized technology capable of discriminating enzyme interactions that occur within complex, biological environments. In a recent publication by Lovell et al, the kallikrein-related peptidase (KLK) family of serine proteases that is highly implicated in the progression of prostate cancer (PCa) was subject to ABPP to elucidate enzymatic activities in the presence of enzalutamide. This is the first report of ABPP in PCa and of activity-based chemical probes selective for individual KLKs. Further, the study reveals androgen receptor-dependent activity among KLK proteins, particularly in mediating the invasion of the bone microenvironment.

Kallikrein-related peptidase (KLK)6 is associated with inflammatory diseases and neoplastic progression. KLK6 is aberrantly expressed in several solid tumors and regulates cancer development, metastatic progression, and drug resistance. However, the function of KLK6 in the tumor microenvironment remains unclear. This study aimed to determine the role of KLK6 in the tumor microenvironment. Here, we uncovered the mechanism underlying KLK6-mediated cross-talk between cancer cells and macrophages. Compared with wild-type mice, KLK6-/- mice showed less tumor growth and metastasis in the B16F10 melanoma and Lewis lung carcinoma (LLC) xenograft model. Mechanistically, KLK6 promoted the secretion of tumor necrosis factor-alpha (TNF-α) from macrophages via the activation of protease-activated receptor-1 (PAR1) in an autocrine manner. TNF-α secreted from macrophages induced the release of the C-X-C motif chemokine ligand 1 (CXCL1) from melanoma and lung carcinoma cells in a paracrine manner. The introduction of recombinant KLK6 protein in KLK6-/- mice rescued the production of TNF-α and CXCL1, tumor growth, and metastasis. Inhibition of PAR1 activity suppressed these malignant phenotypes rescued by rKLK6 in vitro and in vivo. Our findings suggest that KLK6 functions as an important molecular link between macrophages and cancer cells during malignant progression, thereby providing opportunities for therapeutic intervention., Competing Interests: The authors declare no conflicts of interest.

The epidermis is a barrier that prevents water loss while keeping harmful substances from penetrating the host. The impermeable cornified layer of the stratum corneum is maintained by balancing continuous turnover driven by epidermal basal cell proliferation, suprabasal cell differentiation, and corneal shedding. The epidermal desquamation process is tightly regulated by balance of the activities of serine proteases of the Kallikrein-related peptidases (KLK) family and their cognate inhibitor lymphoepithelial Kazal type-related inhibitor (LEKTI), which is encoded by the serine peptidase inhibitor Kazal type 5 gene. Imbalance of proteolytic activity caused by a deficiency of LEKTI leads to excessive desquamation due to increased activities of KLK5, KLK7, and KLK14 and results in Netherton syndrome (NS), a debilitating condition with an unmet clinical need. Increased activity of KLKs may also be pathological in other dermatoses such as atopic dermatitis (AD). Here, we describe the discovery of inhibitory antibodies against murine KLK5 and KLK7 that could compensate for the deficiency of LEKTI in NS. These antibodies are protective in mouse models of NS and AD and, when combined, promote improved skin barrier integrity and reduced inflammation. To translate these findings, we engineered a humanized bispecific antibody capable of potent inhibition of human KLK5 and KLK7. A crystal structure of KLK5 bound to the inhibitory Fab revealed that the antibody binds distal to its active site and uses a relatively unappreciated allosteric inhibition mechanism. Treatment with the bispecific anti-KLK5/7 antibody represents a promising therapy for clinical development in NS and other inflammatory dermatoses.

Pancreatic cancer has the lowest survival rate of all common cancers due to late diagnosis and limited treatment options. Serine hydrolases are known to mediate cancer progression and metastasis through initiation of signaling cascades and cleavage of extracellular matrix proteins, and the kallikrein-related peptidase (KLK) family of secreted serine proteases have emerging roles in pancreatic ductal adenocarcinoma (PDAC). However, the lack of reliable activity-based probes (ABPs) to profile KLK activity has hindered progress in validation of these enzymes as potential targets or biomarkers. Here, we developed potent and selective ABPs for KLK6 by using a positional scanning combinatorial substrate library and characterized their binding mode and interactions by X-ray crystallography. The optimized KLK6 probe IMP-2352 ( k obs / I = 11,000 M -1 s -1 ) enabled selective detection of KLK6 activity in a variety of PDAC cell lines, and we observed that KLK6 inhibition reduced the invasiveness of PDAC cells that secrete active KLK6. KLK6 inhibitors were combined with N-terminomics to identify potential secreted protein substrates of KLK6 in PDAC cells, providing insights into KLK6-mediated invasion pathways. These novel KLK6 ABPs offer a toolset to validate KLK6 and associated signaling partners as targets or biomarkers across a range of diseases.

Estradiol-17β (E2) increases kallikrein in rodent and human reproductive tissues. Kallikrein specific activity is increased in the porcine uterus when conceptus E2 is secreted at maternal recognition of pregnancy. When kallikrein acts on kininogen to liberate bradykinin, angiogenic and vasoactive factors are released. The uterus of ovariectomized ewes administered E2 undergoes rapid vascular changes via different patterns of angiogenic and vasoactive factors. Our hypothesis was that E2 would increase the specific activity and protein secretion of tissue kallikrein in endometrial explants culture media (ECM) and ewes exposed to E2 would have uterine arteries that would be more sensitive to the vasodilatory effects of bradykinin. Ovariectomized ewes received 100 mg of E2 implants for 0, 12, 24, or 48 h. After treatment, uterine weights were determined, and caruncles were processed for ECM. Uterine weights and uterine weight per ewe body weight were significantly greater in the 12 and 24 h ewes compared with the 0 h ewes, with the 48 h ewes being similar to the 24 h ewes. There were no statistically significant differences in caruncular tissue kallikrein protein secretion among the treatment groups. There was a tendency (P = 0.09) for duration of E2 exposure to influence tissue kallikrein specific activity where kallikrein activity was greater (P ≤ 0.05) in the 12 and 48 h ewes compared with the 0 h ewes, with 24 h ewes being intermediate (unprotected F test). Uterine arteries from ewes with E2 for 24 and 48 h had more sensitivity to bradykinin, via the bradykinin receptor 2, than uterine arteries from ewes with 0 or 12 h E2 exposure. We fail to reject our hypothesis as E2 did elicit a positive response in tissue kallikrein specific activity and bradykinin response. Further investigations are needed to determine how kallikrein and bradykinin may be involved in vascular remodeling of the ovine uterus., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: Clinical studies of type 2 (T2) cytokine-related neutralizing antibodies in asthma have identified a substantial subset of patients with low levels of T2 inflammation who do not benefit from T2 cytokine neutralizing antibody treatment. Non-T2 mechanisms are poorly understood in asthma but represent a redefined unmet medical need., Objective: We sought to gain a better understanding of genetic contributions to T2-low asthma., Methods: We utilized an unbiased genome-wide association study of patients with moderate to severe asthma stratified by T2 serum biomarker periostin. We also performed additional expression and biological analysis for the top genetic hits., Results: We identified a novel protective single nucleotide polymorphism at chr19q13.41, which is selectively associated with T2-low asthma and establishes Kallikrein-related peptidase 5 (KLK5) as the causal gene mediating this association. Heterozygous carriers of the single nucleotide polymorphisms have reduced KLK5 expression. KLK5 is secreted by human bronchial epithelial cells and elevated in asthma bronchial alveolar lavage. T2 cytokines IL-4 and IL-13 downregulate KLK5 in human bronchial epithelial cells. KLK5, dependent on its catalytic function, induces epithelial chemokine/cytokine expression. Finally, overexpression of KLK5 in airway or lack of an endogenous KLK5 inhibitor, SPINK5, leads to spontaneous airway neutrophilic inflammation., Conclusion: Our data identify KLK5 to be the causal gene at a novel locus at chr19q13.41 associated with T2-low asthma., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Various approaches have been explored to study skin biology, including the use of stem cells. Mesenchymal stem cells (MSCs) from the umbilical cord can be safely and easily obtained; however, a simple strategy to monitor their differentiation is essential. Involucrin is a marker of keratinocyte differentiation, and its promoter (pINV) directs stratum-specific expression of this protein. We designed a reporter system containing EGFP under the control of pINV to assess MSC transdifferentiation into keratinocytes. The functional sequence of pINV was inserted into a lentiviral vector, producing LeGO-GpINV. MSCs were transduced with LeGO-GpINV and induced to transdifferentiate into keratinocytes under cultivation with keratinocyte serum-free medium. MSC transdifferentiation was confirmed by morphological changes and by the expression of epidermal markers by flow cytometry, quantitative PCR, Western blot and the activity of epidermal kallikreins 5, 6 and 7. After 14 days of transdifferentiation, MSCs transduced with LeGO-GpINV showed an increase in EGFP fluorescence and expressed CK10, CK14, involucrin and filaggrin. There was also an increase in kallikrein activity. This reporter system allowed us to temporally assess epidermal differentiation, simultaneously with involucrin expression, opening possibilities for the in vivo study of skin biology and in regenerative medicine., (Copyright © 2022 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

The epidermis is the outermost skin layer and is part of one of the largest organs in the body; it is supported by the dermis, a network of fibrils, blood vessels, pilosebaceous units, sweat glands, nerves, and cells. The skin as a whole is a protective shield against numerous noxious agents, including microorganisms and chemical and physical factors. These functions rely on the activity of multiple growth factors, peptide hormones, proteases, and specific signaling pathways that are triggered by the activation of distinct types of receptors sited in the cell membranes of the various cell types present in the skin. The human kallikrein family comprises a large group of 15 serine proteases synthesized and secreted by different types of epithelial cells throughout the body, including the skin. At this site, they initiate a proteolytic cascade that generates the active forms of the proteases, some of which regulate skin desquamation, activation of cytokines, and antimicrobial peptides. Kinin peptides are formed by the action of plasma and tissue kallikreins on kininogens, two plasma proteins produced in the liver and other organs. Although kinins are well known for their proinflammatory abilities, in the skin they are also considered important modulators of keratinocyte differentiation. In this review, we summarize the contributions of the kallikreins and kallikrein-related peptidases family and those of kinins and their receptors in skin homeostasis, with special emphasis on their pathophysiological role.

Right ventricular (RV) hypertrophy and further heart failure are major co-morbidities, resulting in the premature death of patients with hypoxic pulmonary hypertension (HPH). The regulatory effects of kallikrein-related peptidase (KLK) family members on cardiac function have been extensively studied. However, to the best of the authors' knowledge, the regulatory effects of KLK8 on RV hypertrophy caused by HPH have yet to be reported. The aim of the present study was to assess KLK8 expression in the RV tissue of HPH-modeled rats, and to further explore the effects and underlying mechanism of KLK8 in regulating the hypertrophy of hypoxia-induced H9c2 cardiomyocytes. In HPH model rats, increases in the right ventricle hypertrophy index, the right ventricular systolic pressure, cardiac output, as well as pulmonary artery wall thickness were observed. Western blot analysis revealed that KLK8 expression and MAPK/p53 signaling activity were enhanced in the RVs of rats in an RV HPH rat model. In hypoxia-induced H9c2 cardiomyocytes, KLK8 overexpression promoted cardiomyocyte hypertrophy, whereas KLK8 silencing showed the opposite results. KLK8 overexpression increased the expression levels of ventricular hypertrophy markers, including atrial natriuretic peptide, brain natriuretic peptide and myosin heavy chain 7, which were blocked upon addition of the p38 MAPK inhibitor, SB202190. Conversely, KLK8 silencing caused a decrease in the expression levels of the ventricular hypertrophy markers, which were further reduced via inhibition of the p38 MAPK/p53 signaling pathway. Taken together, the results of the present study have shown that KLK8 may subtly regulate RV hypertrophy, and therefore KLK8 may be a promising therapeutic target for treating HPH-induced RV hypertrophy., (Copyright © 2022 Elsevier Ltd. All rights reserved.)