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RNA therapeutics hold significant promise in the treatment of cardiovascular diseases. RNAs are biologically diverse and functionally specific and can be used for gain- or loss-of-function purposes. The effectiveness of mRNA-based vaccines in the recent COVID-19 pandemic has undoubtedly proven the benefits of an RNA-based approach. RNA-based therapies are becoming more common as a treatment modality for cardiovascular disease. This is most evident in hypertension where several small interfering RNA–based drugs have proven to be effective in managing high blood pressure in several clinical trials. As befits a rapidly burgeoning field, there is significant interest in other classes of RNA. Revascularization of the infarcted heart through an mRNA drug is under clinical investigation. mRNA technology may provide the platform for the expression of paracrine factors for myocardial protection and regeneration. Emergent technologies on the basis of microRNAs and gene editing are tackling complex diseases in a novel fashion. RNA-based gene editing offers hope of permanent cures for monogenic cardiovascular diseases, and long-term control of complex diseases such as essential hypertension, as well. Likewise, microRNAs are proving effective in regenerating cardiac muscle. The aim of this review is to provide an overview of the current landscape of RNA-based therapies for the treatment of cardiovascular disease. The review describes the large number of RNA molecules that exist with a discussion of the clinical development of each RNA type. In addition, the review also presents a number of avenues for future development. [ABSTRACT FROM AUTHOR]

Gynecologic tumors are among the leading diseases in women in terms of morbidity and mortality, and patients with recurrence and drug resistance after conventional treatment have poor prognosis. Small interfering RNA (siRNA) is a promising tool for the treatment of gene-related diseases because of its high efficiency in gene silencing, ease of synthesis and wide range of targets. However, the clinical application of siRNA drugs still needs to face the problems of bare sequence instability, low in vivo delivery efficiency, and uncontrollable adverse effects. In recent years, the choice of vectors for in vivo delivery of siRNA drugs and the selection of suitable targets for their application have been the focus of research on siRNA therapeutics. This paper mainly reviews the recent research progress about the application of vector-modified siRNA drugs in gynecologic tumor therapy and the important potential application targets of siRNA in various gynecologic tumors. [ABSTRACT FROM AUTHOR]

PIWI-interacting RNA (piRNA), a kind of non-coding RNAs with a length of about 24-31 nucleotides, is specifically expressed in animal germ cells. The production process of piRNA can be divided into two parts: transcription in the nucleus and post-processing in the cytoplasm. piRNA can silence transcriptional genes, and regulates mRNA with the help of PIWI family proteins, and further regulates reproductive function. In male reproductive function, piRNA can control spermatogenesis by RNF8, glutathione peroxidase 5 and other proteins with the help of PIWI family proteins, resulting in azoospermia, oligozoospermia, asthenospermia and other diseases affecting male reproductive function. In the initial study of piRNA, researcher generally believed that piRNA had little effect on female reproduction. However, recent studies on golden hamsters found that piRNA/ PIWI also played an important role in the regulation of female reproductive function. The production of piRNA and its correlation with reproductive function were reviewed. [ABSTRACT FROM AUTHOR]

The poor prognosis of hepatocellular carcinoma (HCC) is still an urgent challenge to be solved worldwide. Hence, assembling drugs and targeted short peptides together to construct a novel medicine delivery strategy is crucial for targeted and synergy therapy of HCC. Herein, a high-efficiency nanomedicine delivery strategy has been constructed by combining graphdiyne oxide (GDYO) as a drug-loaded platform, specific peptide (SP94-PEG) as a spear to target HCC cells, sorafenib, doxorubicin-Fe 2+ (DOX-Fe 2+ ), and siRNA (SLC7A11-i) as weapons to exert a three-path synergistic attack against HCC cells. In this work, SP94-PEG and GDYO form nanosheets with HCC-targeting properties, the chemotherapeutic drug DOX linked to ferrous ions increases the free iron pool in HCC cells and synergizes with sorafenib to induce cell ferroptosis. As a key gene of ferroptosis, interference with the expression of SLC7A11 makes the ferroptosis effect in HCC cells easier, stronger, and more durable. Through gene interference, drug synergy, and short peptide targeting, the toxic side effects of chemotherapy drugs are reduced. The multifunctional nanomedicine GDYO@SP94/DOX-Fe 2+ /sorafenib/SLC7A11-i (MNMG) possesses the advantages of strong targeting, good stability, the ability to continuously induce tumor cell ferroptosis and has potential clinical application value, which is different from traditional drugs., (© 2024. The Author(s).)

Competing Interests: Conflict of interest: Professor Webb’s employer, the University of Edinburgh, has been paid by Alnylam for his contributions to the zilebesiran programme in hypertension. He has undertaken recent consultancy roles with Idorsia, Janssen, and Silence pharmaceuticals in the field of cardiovascular research.

Over the last decade, randomized clinical trials of several pharmacologic agents have demonstrated a reduction in cardiovascular mortality and other important secondary outcomes. Angiotensin-Neprilysin Inhibitors and Sodium-Glucose Co-transporter 2 inhibitors have now become pillars in the treatment of heart failure. Ivabradine is a negative chronotropic agent used as an adjunctive therapy in patients with heart failure. Two new hypertension therapies, zilebresiran and aprocitentan, are currently in investigational stages. Finally, mavacamten has emerged as a pharmacologic treatment for hypertrophic obstructive cardiomyopathy. Practitioners must be familiar with the indications and side effects of newer therapies as they are now frequently prescribed., Competing Interests: Disclosure The authors have nothing to disclose., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background and Purpose: Hereditary transthyretin-mediated amyloidosis with polyneuropathy (ATTRv-PN [v for variant]) is a rare, progressive disease associated with multisystemic impairments. This study assessed the real-world outcomes of patients with ATTRv-PN who switched from tafamidis to patisiran, as well as the reasons for the treatment switch., Methods: This was a retrospective chart review study at a large expert referral center. Data were extracted from medical charts of patients with ATTRv-PN who switched from tafamidis to patisiran on or before 30 August 2019. Data elements included demographic and clinical characteristics, rationale for switch, and disease measures evaluated from tafamidis initiation through the 12-month patisiran treatment period., Results: Among the 24 patients with ATTRv-PN included in the study, 50.0% had a V30M variant, and the mean (SD) age was 67.3 (8.0) years. During tafamidis treatment (mean [SD] = 30.1 [17.5] months) before switching to patisiran, patients worsened across multiple polyneuropathy measures, including walking ability, Neuropathy Impairment Score, and autonomic function. Neuropathic disease progression on tafamidis was the principal reason for switching to patisiran. After 12 months on patisiran (mean [SD] = 11.7 [1.4] months), patients experienced attenuated disease progression or improvement in the aforementioned measures of polyneuropathy., Conclusions: Switching from tafamidis to patisiran attenuated the rate of functional decline, and most patients experienced stabilization or improvement of at least one polyneuropathy measure within 12 months of patisiran treatment. Timely switch from tafamidis to patisiran can be beneficial to avoid rapid disease progression in patients with ATTRv-PN., (© 2024 Analysis Group, Alnylam Pharmaceuticals and The Authors. European Journal of Neurology published by John Wiley & Sons Ltd on behalf of European Academy of Neurology.)

Background: Elevated levels of lipoprotein(a) [Lp(a)] represent a risk factor for cardiovascular disease including aortic valve stenosis, myocardial infarction and stroke. While the patho-physiological mechanisms linking Lp(a) with atherosclerosis are not fully understood, from genetic studies that lower Lp(a) levels protect from CVD independently of other risk factors including lipids and lipoproteins. Hereby, Lp(a) has been considered an appealing pharmacological target., Results: However, approved lipid lowering therapies such as statins, ezetimibe or PCSK9 inhibitors have a neutral to modest effect on Lp(a) levels, thus prompting the development of new strategies selectively targeting Lp(a). These include antisense oligonucleotides and small interfering RNAs (siRNAs) directed towards apolipoprotein(a) [Apo(a)], which are in advanced phase of clinical development. More recently, additional approaches including inhibitors of Apo(a) and gene editing approaches via CRISPR-Cas9 technology entered early clinical development., Conclusion: If the results from the cardiovascular outcome trials, designed to demonstrate whether the reduction of Lp(a) of more than 80% as observed with pelacarsen, olpasiran or lepodisiran translates into the decrease of cardiovascular mortality and major adverse cardiovascular events, will be positive, lowering Lp(a) will become a new additional target in the management of patients with elevated cardiovascular risk., (© 2024 Stichting European Society for Clinical Investigation Journal Foundation. Published by John Wiley & Sons Ltd.)

Recent single-cell RNA-sequencing analysis of rheumatoid arthritis (RA) synovial tissues revealed the heterogeneity of RA synovial fibroblasts (SFs) with distinct functions such as high IL-6 production. The molecular mechanisms responsible for high IL-6 production will become a promising drug target of RASFs to treat RA. In this study, we performed siRNA screening of 65 transcription factors (TFs) differentially expressed among RASF subsets to identify TFs involved in IL-6 production. The siRNA screening identified 7 TFs including ARID5B , a RA risk gene, that affected IL-6 production. Both long and short isoforms of ARID5B were expressed and negatively regulated by TNF-α in RASFs. The siRNA knockdown and lentiviral overexpression of long and short isoforms of ARID5B revealed that the long isoform suppressed IL-6 production stimulated with TNF-α. eQTL analysis using 58 SFs demonstrated that RA risk allele, rs10821944, in intron 4 of the ARID5B gene had a trend of eQTL effects to the expression of long isoform of ARID5B in SFs treated with TNF-α. ARID5B was found to be a negative modulator of IL-6 production in RASFs. The RA risk allele of ARID5B intron may cause high IL-6 production, suggesting that ARID5B will become a promising drug target to treat RA.

Purpose of Review: Tafamidis is currently the only approved disease-modifying treatment for ATTR-CM. However, there have been important developments in the treatment of ATTR-CM, as the results of two phase 3 trials were published and several other trials are in their final stages. In this review, we summarize current and future therapies for ATTR-CM., Recent Findings: Recently, acoramidis, a TTR stabilizer has been proven to be effective in reducing mortality and morbidity compared to placebo in the ATTRibute-CM trial. Additionally, patisiran, an RNA silencer, preserved functional capacity and quality of life compared to placebo in the APOLLO-B trial. However, the FDA declined to approve patisiran for ATTR-CM. The results of phase 1 trial of ALXN2220, an antiamyloid antibody raise hope for reversal of myocardial damage by amyloid depletion. Phase 3 trials evaluating the efficacy of different RNA silencers, gene editing with CRISPR-Cas9, and other anti-amyloid antibodies are ongoing., Summary: Therapies targeting different mechanism in the pathophysiology of ATTR-CM provide new alternatives for treating patients with ATTR-CM. Future research should focus on comparing their effectiveness, the potential of combined treatment with agents from different classes and on identifying the patients who will benefit most from each class of medication., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Methotrexate (MTX) is commonly prescribed as the initial treatment for gestational trophoblastic neoplasia (GTN), but MTX monotherapy may not be effective for high-risk GTN and choriocarcinoma. The cellular uptake of MTX is essential for its pharmacological activity. Thus, our study aimed to investigate the cellular pharmacokinetics and transport mechanisms of MTX in choriocarcinoma cells. For the quantification of MTX concentrations in cellular matrix, a liquid chromatography-tandem mass spectrometry method was created and confirmed initially. MTX accumulation in BeWo, JEG-3, and JAR cells was minimal. Additionally, the mRNA levels of folate receptor α (FRα) and breast cancer resistance protein (BCRP) were relatively high in the three choriocarcinoma cell lines, whereas proton-coupled folate transporter (PCFT), reduced folate carrier (RFC), and organic anion transporter (OAT) 4 were low. Furthermore, the expression of other transporters was either very low or undetectable. Notably, the application of inhibitors and small interfering RNAs (siRNAs) targeting FRα, RFC, and PCFT led to a notable decrease in the accumulation of MTX in BeWo cells. Conversely, the co-administration of multidrug resistance protein 1 (MDR1) and BCRP inhibitors increased MTX accumulation. In addition, inhibitors of OATs and organic-anion transporting polypeptides (OATPs) reduced MTX accumulation, while peptide transporter inhibitors had no effect. Results from siRNA knockdown experiments and transporter overexpression cell models indicated that MTX was not a substrate of nucleoside transporters. In conclusion, the results indicate that FRα and multiple transporters such as PCFT, RFC, OAT4, and OATPs are likely involved in the uptake of MTX, whereas MDR1 and BCRP are implicated in the efflux of MTX from choriocarcinoma cells. These results have implications for predicting transporter-mediated drug interactions and offer potential directions for further research on enhancing MTX sensitivity., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Mengru Bai reports financial support was provided by National Natural Science Foundation of China. Mengru Bai reports financial support was provided by Zhejiang Province Natural Science Foundation of China. Yong Wu reports financial support was provided by Zhejiang Provincial Medicine and Health Science Foundation. If there are other authors, they 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.)

Endotracheal Tubes (ETTs) maintain and secure a patent airway; however, prolonged intubation often results in unintended injury to the mucosal epithelium and inflammatory sequelae which complicate recovery. ETT design and materials used have yet to adapt to address intubation associated complications. In this study, a composite coating of electrospun polycaprolactone (PCL) fibers embedded in a four-arm polyethylene glycol acrylate matrix (4APEGA) is developed to transform the ETT from a mechanical device to a dual-purpose device capable of delivering multiple therapeutics while preserving coating integrity. Further, the composite coating system (PCL-4APEGA) is capable of sustained delivery of dexamethasone from the PCL phase and small interfering RNA (siRNA) containing polyplexes from the 4APEGA phase. The siRNA is released rapidly and targets smad3 for immediate reduction in pro-fibrotic transforming growth factor-beta 1 (TGFϐ1) signaling in the upper airway mucosa as well as suppressing long-term sequelae in inflammation from prolonged intubation. A bioreactor was used to study mucosal adhesion to the composite PCL-4APEGA coated ETTs and investigate continued mucus secretory function in ex vivo epithelial samples. The addition of the 4APEGA coating and siRNA delivery to the dexamethasone delivery was then evaluated in a swine model of intubation injury and observed to restore mechanical function of the vocal folds and maintain epithelial thickness when observed over 14 days of intubation. This study demonstrated that increase in surface lubrication paired with surface stiffness reduction significantly decreased fibrotic behavior while reducing epithelial adhesion and abrasion., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Teja Guda reports financial support was provided by 59MDW Office of the Chief Scientist. Teja Guda has patent pending to UT System Board of Regents., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by joint inflammation and destruction. Current treatments, such as Methotrexate (MTX), though effective, often face limitations such as high plasma C max and lack of sustained release. This study explores a synergistic approach to RA therapy using folate-liposomal co-delivery of MTX and RELA siRNA (short interfering RNA), targeting RAW264.7 macrophage repolarization via nuclear factor kappa B (NF-κB) pathway inhibition. Extensive in vitro characterizations demonstrate the stability and biocompatibility of this therapy via folate-liposomes. In the collagen-induced arthritis (CIA) rat model, treatment leads to reduced synovial inflammation and improved mobility. The combined MTX and RELA siRNA approach indirectly inhibits inflammatory cytokines, rheumatoid factor (RF), and C-reactive protein (CRP). Targeted macrophage delivery shows marked therapeutic effects in RAW264.7 murine macrophages, potentially modulating M1 to M2 polarization. This research presents a promising avenue for innovative RA therapies by inhibiting the inflammatory cascade and preventing joint damage., (© 2024 Wiley‐VCH GmbH.)

Sympathetic nerves play a pivotal role in promoting tumor growth through crosstalk with tumor and stromal cells. Chemotherapy exacerbates the infiltration of sympathetic nerves into tumors, thereby providing a rationale for inhibiting sympathetic innervation to enhance chemotherapy. Here, we discovered that doxorubicin increases the density and activity of sympathetic nerves in breast cancer mainly by upregulating the expression of nerve growth factors (NGFs) in cancer cells. To address this, we developed a combination therapy by co-encapsulating small interfering RNA (siRNA) and doxorubicin within breast cancer-targeted poly (lactic-co-glycolic acid) (PLGA) nanoparticles, aiming to suppress NGF expression post-chemotherapy. Incorporating NGF blockade into the nanoplatform for chemotherapy effectively mitigated the chemotherapy-induced proliferation of sympathetic nerves. This not only bolstered the tumoricidal activity of chemotherapy, but also amplified its stimulatory impact on the antitumor immune response by increasing the infiltration of immunostimulatory cells into tumors while concurrently reducing the frequency of immunosuppressive cells. Consequently, the combined nanodrug approach, when coupled with anti-PD-L1 treatment, exhibited a remarkable suppression of primary and deeply metastatic tumors with minimal systematic toxicity. Importantly, the nanoplatform relieved chemotherapy-induced peripheral neuropathic pain (CIPNP) by diminishing the expression of pain mediator NGFs. In summary, this research underscores the significant potential of NGF knockdown in enhancing immunochemotherapy outcomes and presents a nanoplatform for the highly efficient and low-toxicity treatment of breast cancer., 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 Ltd. All rights reserved.)

Objectives: Airway remodeling, a prominent feature of asthma, involves aberrant proliferation, apoptosis, and migration of airway smooth muscle cells (ASMCs). Toll-like receptors (TLRs) are implicated in the regulation of the autophagy pathway. In this study, we aimed to investigate the influence of Toll-like receptor 4 (TLR4) on autophagy and its underlying mechanism in ASMC proliferation, apoptosis, and migration., Methods: Histopathological changes in the lungs of asthmatic mice assessed by Hematoxylin-Eosin (HE) and Masson staining. Cell proliferation, apoptosis and migration were evaluated utilizing CCK8, Edu, Flow cytometry and wound heading assays. The effectiveness of siRNA transfection and the expression of TLR4, autophagy, and proliferation-related proteins after siRNA treatment were examined through RT-PCR and Western blot (WB)., Conclusion: We observed an increase in TLR4 expression and autophagy in a mouse model of OVA-induced asthma. In vitro experiments showed that siRNA-mediated inhibition of TLR4 suppressed autophagy, proliferation, and migration of ASMCs, whereas TLR4 activation by lipopolysaccharide (LPS) had the opposite effect. Furthermore, the autophagy inhibitor 3-Methyladenine (3MA) inhibited ASMCs proliferation and migration while promoting apoptosis. Significantly, our study demonstrated that autophagy inhibition reversed the promotion effect of LPS on ASMC proliferation and migration. These findings suggest that TLR4 may modulate ASMC behavior through the regulation of autophagy.

Background: The Hippo pathway is a conserved tumour suppressor signalling pathway, and its dysregulation is often associated with abnormal cell growth and tumorigenesis. We previously revealed that the transcriptional coactivator Yes-associated protein (YAP), the key effector of the Hippo pathway, is a molecular target for glioblastoma (GBM), the most common malignant brain tumour. Inhibiting YAP with small interfering RNA (siYAP) or the specific inhibitor verteporfin (VP) can diminish GBM growth to a certain degree., Results: In this study, to enhance the anti-GBM effect of siYAP and VP, we designed stepwise-targeting and hypoxia-responsive liposomes (AMVY@NPs), which encapsulate hypoxia-responsive polymetronidazole-coated VP and DOTAP adsorbed siYAP, with angiopep-2 (A2) modification on the surface. AMVY@NPs exhibited excellent blood‒brain barrier crossing, GBM targeting, and hypoxia-responsive and efficient siYAP and VP release properties. By inhibiting the expression and function of YAP, AMVY@NPs synergistically inhibited both the growth and stemness of GBM in vitro. Moreover, AMVY@NPs strongly inhibited the growth of orthotopic U87 xenografts and improved the survival of tumour-bearing mice without adverse effects., Conclusion: Specific targeting of YAP with stepwise-targeting and hypoxia-responsive liposome AMVY@NPs carrying siYAP and VP efficiently inhibited GBM progression. This study provides a valuable drug delivery platform and creative insights for molecular targeted treatment of GBM in the future., (© 2024. The Author(s).)

Abdominal aortic aneurysm (AAA) is a life-threatening vascular disease, while there is a lack of pharmaceutical interventions to halt AAA progression presently. To address the multifaceted pathology of AAA, this work develops a novel multifunctional gene delivery system to simultaneously deliver two siRNAs targeting MMP-2 and MMP-9. The system (TPNs-siRNA), formed through the oxidative polymerization and self-assembly of epigallocatechin gallate (EGCG), efficiently encapsulates siRNAs during self-assembly. TPNs-siRNA safeguards siRNAs from biological degradation, facilitates intracellular siRNA transfection, promotes lysosomal escape, and releases siRNAs to silence MMP-2 and MMP-9. Additionally, TPNs, serving as a multi-bioactive material, mitigates oxidative stress and inflammation, fosters M1-to-M2 repolarization of macrophages, and inhibits cell calcification and apoptosis. In experiments with AAA mice, TPNs-siRNA accumulated and persisted in aneurysmal tissue after intravenous delivery, demonstrating that TPNs-siRNA can be significantly distributed in macrophages and VSMCs relevant to AAA pathogenesis. Leveraging the carrier's intrinsic multi-bioactive properties, the targeted siRNA delivery by TPNs exhibits a synergistic effect for enhanced AAA therapy. Furthermore, TPNs-siRNA is gradually metabolized and excreted from the body, resulting in excellent biocompatibility. Consequently, TPNs emerges as a promising multi-bioactive nanotherapy and a targeted delivery nanocarrier for effective AAA therapy., (© 2024. The Author(s).)

Rising global populations have amplified food scarcity and ushered in the development of genetically modified (GM) crops containing small interference RNAs (siRNAs) that control gene expression to overcome these challenges. The use of RNA interference (RNAi) in agriculture remains controversial due to uncertainty regarding the unintended release of genetic material and downstream nontarget effects, which have not been assessed in environmental bacteria to date. To evaluate the impacts of siRNAs used in agriculture on environmental bacteria, this study assessed microbial growth and viability as well as transcription activity with and without the presence of environmental stressors. Results showed a statistically significant reduction in growth capacity and maximum biomass achieved when bacteria are exposed to siRNAs alone and with additional external stress ( p < 0.05). Further transcriptomic analysis demonstrated that nutrient cycling gene activities were found to be consistently and significantly altered following siRNA exposure, particularly among carbon ( xylA, FBPase , limEH, Chitinase, rgl, rgh, rgaE, mannanase, ara ) and nitrogen ( ureC, nasA, narB, narG, nirK ) cycling genes ( p < 0.05). Decreases in carbon cycling gene transcription profiles were generally significantly enhanced when siRNA exposure was coupled with nutrient or antimicrobial stress. Collectively, findings suggest that certain conditions facilitate the uptake of siRNAs from their surrounding environments that can negatively affect bacterial growth and gene expression activity, with uncertain downstream impacts on ecosystem homeostasis.

PM 2.5 pollution has been associated with the incidence of lung cancer, but the underlying mechanism is still unclear. PIWI-interacting RNAs (piRNAs), initially identified in germline cells, have emerged as a novel class of small non-coding RNAs (26 - 32 nucleotides) with diverse functions in various diseases, including cancer. However, the role and mechanism of piRNAs in the development of PM 2.5 -induced lung cancer remain to be clarified. In the presented study, we used a PM 2.5 -induced malignant transformation cell model to analyze the change of piRNA profiles. Among the disturbed piRNAs, piR-27222 was identified as an oncogene that inhibited cell death in a m 6 A-dependent manner. Mechanistically, we found that piR-27222 could deubiquitinate and stabilize eIF4B by directly binding to eIF4B and reducing its interaction with PARK2. The enhanced expression of eIF4B, in turn, promoted the expression of WTAP, leading to increased m 6 A modification in the Casp8 transcript. Consequently, the stability of Casp8 transcripts was reduced, rendering lung cancer cells resistant to PANoptosis. Collectively, our findings reveal that PM 2.5 exposure up-regulated piR-27222 expression, which could affect EIF4B/WTAP/m 6 A axis, thereby inhibiting PANoptosis of cells and promoting lung cancer. Our study provides new insights into understanding the epigenetic mechanisms underlining PM 2.5 -induced lung cancer., 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 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: CaM (calmodulin)-mediated long-QT syndrome is a genetic arrhythmia disorder (calmodulinopathies) characterized by a high prevalence of life-threatening ventricular arrhythmias occurring early in life. Three distinct genes ( CALM1 , CALM2 , and CALM3 ) encode for the identical CaM protein. Conventional pharmacotherapies fail to adequately protect against potentially lethal cardiac events in patients with calmodulinopathy., Methods: Five custom-designed CALM1 -, CALM2 -, and CALM3 -targeting short hairpin RNAs (shRNAs) were tested for knockdown (KD) efficiency using TSA201 cells and reverse transcription-quantitative polymerase chain reaction. A dual-component suppression and replacement (SupRep) CALM gene therapy (CALM-SupRep) was created by cloning into a single construct CALM1 -, CALM2 -, and CALM3-specific shRNAs that produce KD (suppression) of each respective gene and a shRNA-immune CALM1 cDNA (replacement). CALM1-F142L, CALM2-D130G, and CALM3-D130G induced pluripotent stem cell-derived CMs were generated from patients with CaM-mediated long-QT syndrome. A voltage-sensing dye was used to measure action potential duration at 90% repolarization (APD90)., Results: Following shRNA KD efficiency testing, a candidate shRNA was identified for CALM1 (86% KD), CALM2 (71% KD), and CALM3 (94% KD). The APD90 was significantly prolonged in CALM2-D130G (647±9 ms) compared with CALM2-WT (359±12 ms; P <0.0001). Transfection with CALM -SupRep shortened the average APD90 of CALM2-D130G to 457±19 ms (66% attenuation; P <0.0001). Additionally, transfection with CALM -SupRep shortened the APD90 of CALM1-F142L (665±9 to 410±15 ms; P <0.0001) and CALM3-D130G (978±81 to 446±6 ms; P <0.001)., Conclusions: We provide the first proof-of-principle suppression-replacement gene therapy for CaM-mediated long-QT syndrome. The CALM-SupRep gene therapy shortened the pathologically prolonged APD90 in CALM1 -, CALM2 -, and CALM3-variant CaM-mediated long-QT syndrome induced pluripotent stem cell-derived CM lines. The single CALM-SupRep construct may be able to treat all calmodulinopathies, regardless of which of the 3 CaM-encoding genes are affected., Competing Interests: Dr Ackerman is a consultant for Abbott, Boston Scientific, Bristol Myers Squibb, Daiichi Sankyo, Invitae, Medtronic, Tenaya Therapeutics, and UpToDate. Dr Ackerman and Mayo Clinic are involved in an equity/royalty relationship with AliveCor, Anumana, ARMGO Pharma, Pfizer, and Thryv Therapeutics. However, none of these entities have contributed to this study in any manner. The other authors report no conflicts.

Melatonin, the multi-functional neurohormone, is synthesized in the extra-pineal tissues such as the hippocampus. The key enzyme in hippocampal melatonin synthesis is arylalkylamine-N-acetyltransferase (AANAT). The importance of melatonin synthesis in the hippocampus has not yet been determined. We investigated hippocampal AANAT role in cognitive function using gene silencing small interference RNA (siRNA) technology. The hippocampal local melatonin synthesis was inhibited by AANAT-siRNA injection. The time-gene silencing profile of AANAT-siRNA was obtained by RT-PCR technique. The cytotoxicity of siRNA dose was determined by MTT assay on the B65 neural cells. Animals received the selected dosage of AANAT-siRNA. Then, the spatial working memory (Y maze), object recognition memory and spatial reference memory (Morris's water maze, MWM) were evaluated. The anxiety-like behaviors were evaluated by the elevated plus maze. After one week, following the probe test of MWM, the rats were sacrificed for histological analysis. The hippocampal melatonin levels were measured using the liquid chromatography-mass spectrometry technique. The hippocampal melatonin levels in the AANAT-siRNA group decreased. Animals receiving the AANAT-siRNA showed deficits in spatial learning and working memory which were verified by increased escape latency and reduced spontaneous alternations, respectively. There was an increase in anxiety-like behaviors as well as a deficit in recognition memory in the AANAT-siRNA group. The Nissl staining and immunohistochemistry of activated caspase-3 showed the neuronal loss and cell apoptosis in hippocampal tissue of the AANAT-siRNA group. The 18 F-FDG-PET imaging displayed lower glucose metabolism following the reduction in AANAT mRNA. Data suggest that the AANAT mRNA and hippocampal melatonin synthesis might be an essential factor for learning, memory and some aspects of cognition, as well as homeostasis of hippocampal cells., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Cellular communication among different types of vascular cells is indispensable for maintaining vascular homeostasis and preventing atherosclerosis. However, the biological mechanism involved in cellular communication among these cells and whether this biological mechanism can be used to treat atherosclerosis remain unknown. We hypothesized that endothelial autophagy mediates the cellular communication in vascular tissue through exosome-mediated delivery of atherosclerosis-related genes., Methods: Rapamycin and adeno-associated virus carrying Atg7 short hairpin RNA under the Tie (TEK receptor tyrosine kinase) promoter were used to activate and inhibit vascular endothelial autophagy in high-fat diet-fed ApoE -/- mice, respectively. miRNA microarray, in vivo and in vitro experiments, and human vascular tissue were used to explore the effects of endothelial autophagy on endothelial function and atherosclerosis and its molecular mechanisms. Quantitative polymerase chain reaction and miRNA sequencing were performed to determine changes in miRNA expression in exosomes. Immunofluorescence and exosome coculture experiments were conducted to examine the role of endothelial autophagy in regulating the communication between endothelial cells and smooth muscle cells (SMCs) via exosomal miRNA., Results: Endothelial autophagy was inhibited in thoracic aortas of high-fat diet-fed ApoE -/ - mice. Furthermore, rapamycin alleviated high-fat diet-induced atherosclerotic burden and endothelial dysfunction, while endothelial-specific Atg7 depletion aggravated the atherosclerotic burden. miRNA microarray, in vivo and in vitro experiments, and human vascular tissue analysis revealed that miR-204-5p was significantly increased in endothelial cells after high-fat diet exposure, which directly targeted Bcl2 to regulate endothelial cell apoptosis. Importantly, endothelial autophagy activation decreased excess miR-204-5p by loading miR-204-5p into multivesicular bodies and secreting it through exosomes. Moreover, exosomal miR-204-5p can effectively transport to SMCs, alleviating SMC calcification by regulating target proteins such as RUNX2 (runt-related transcription factor 2)., Conclusions: Our study revealed the exosomal pathway by which endothelial autophagy protects atherosclerosis: endothelial autophagy activation transfers miR-204-5p from endothelial cells to SMCs via exosomes, both preventing endothelial apoptosis and alleviating SMC calcification., Registration: URL: https://www.chictr.org.cn/; Unique identifier: ChiCTR2200064155., Competing Interests: None.

Pathogenic viruses are a profound threat to global public health, underscoring the urgent need for the development of efficacious antiviral therapeutics. The advent of RNA-targeting antiviral strategies has marked a significant paradigm shift in the management of viral infections, offering a potent means of control and potential cure. In this review, we delve into the cutting-edge progress in RNA-targeting antiviral agents, encompassing antisense oligonucleotides (ASOs), small interfering RNAs (siRNAs), small and bifunctional molecules. We provide an in-depth examination of their strategic molecular design and elucidate the underlying mechanisms of action that confer their antiviral efficacy. By synthesizing recent findings, we shed light on the innovative potential of RNA-targeting approaches and their pivotal role in advancing the frontiers of antiviral drug discovery., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Hereditary transthyretin amyloidosis (ATTRv) is a rare, progressive, and life-threatening disease caused by misfolded transthyretin (TTR) proteins that aggregate as abnormal amyloid fibrils and accumulate throughout the body. The kidney is one of the main organs affected in amyloid light chain (AL) amyloidosis and ATTRv amyloidosis. The most common clinical presentation is proteinuria, which consists mainly of albumin; this is the first step in the natural history of ATTRv nephropathy. Not all TTR mutations are equal in terms of ATTRv kidney involvement. Kidney involvement in ATTRv itself is difficult to define, given the numerous associated confounding factors. There are several treatments available to treat ATTRv, including orthotopic liver transplant (OLT), which is the classic treatment for ATTRv. However, we should be careful regarding the use of calcineurin inhibitors in the setting of OLT because these can be nephrotoxic. New treatments for amyloidosis may have an impact on kidney function, including drugs that target specific pathways involved in the disease. Tafamidis and diflunisal, which are TTR stabilizers, patisiran (RNA interference agent), and inotersen (antisense oligonucleotide inhibitor) have been shown to reduce TTR amyloid. Tafamidis and patisiran are medications that have reduced the progression of kidney disease in amyloidosis, but inotersen and diflunisal may damage kidney function., (Copyright © 2024 National Kidney Foundation, Inc. Published by Elsevier Inc. All rights reserved.)

Aims: Excessive occlusal force with periodontitis leads to rapid alveolar bone resorption. However, the molecular mechanism by which inflammation and mechanical stress cause bone resorption remains unclear. We examined the role of Piezo1, a mechanosensitive ion channel expressed on osteoblasts, in the changes in the receptor activator of nuclear factor-kappa B ligand (RANKL)/osteoprotegerin (OPG) ratio in mouse MC3T3-E1 osteoblast-like cells under Porphyromonas gingivalis lipopolysaccharide (P.g.-LPS) and mechanical stress., Methods: To investigate the effect of P.g.-LPS and mechanical stress on the RANKL/OPG ratio and Piezo1 expression, we stimulated MC3T3-E1 cells with P.g.-LPS. After 3 days in culture, shear stress, a form of mechanical stress, was applied to the cells using an orbital shaker. Subsequently, to investigate the role of Piezo1 in the change of RANKL/OPG ratio, we inhibited Piezo1 function by knockdown via Piezo1 siRNA transfection or by adding GsMTx4, a Piezo1 antagonist., Results: The RANKL/OPG ratio significantly increased in MC3T3-E1 cells cultured in a medium containing P.g.-LPS and undergoing mechanical stress compared to cells treated with P.g.-LPS or mechanical stress alone. However, the expression of Piezo1 was not increased by P.g.-LPS and mechanical stress. In addition, phosphorylation of MEK/ERK was induced in the cells under P.g.-LPS and mechanical stress. MC3T3-E1 cells treated with P.g.-LPS and mechanical stress when cocultured with RAW264.7 cells induced their differentiation into osteoclast-like cells. The increased RANKL/OPG ratio was suppressed by either Piezo1 knockdown or the addition of GsMTx4. Furthermore, GsMTx4 inhibited the phosphorylation of MEK/ERK., Conclusion: These findings suggest that P.g.-LPS and Piezo1-mediated mechanical stress induce MEK/ERK phosphorylation and increase RANKL expression in osteoblasts. Consequently, this leads to the differentiation of osteoclast precursor cells into osteoclasts., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Aims: To conduct a pooled analysis of Phase 3 trials investigating the efficacy and safety of inclisiran across glycaemic and body mass index (BMI) strata., Materials and Methods: Participants were randomized 1:1 to receive 300 mg inclisiran sodium or placebo twice yearly, after initial and 3-month doses up to 18 months, with background oral lipid-lowering therapy. Analyses were stratified by glycaemic status (normoglycaemia, prediabetes, and diabetes) or BMI (<25, ≥25 to <30, ≥30 to <35, and ≥35 kg/m 2 ). Co-primary endpoints were percentage and time-adjusted percentage change in low-density lipoprotein (LDL) cholesterol from baseline. Safety was also assessed., Results: Baseline characteristics were balanced between treatment arms and across strata. Percent LDL cholesterol change (placebo-corrected) with inclisiran from baseline to Day 510 ranged from -47.6% to -51.9% and from -48.8% to -54.4% across glycaemic/BMI strata, respectively. Similarly, time-adjusted percentage changes after Day 90 and up to Day 540 ranged from -46.8% to -52.0% and from -48.6% to -53.3% across glycaemic/BMI strata, respectively. Inclisiran led to significant reductions in proprotein convertase subtilisin/kexin type 9 and other atherogenic lipids and lipoproteins versus placebo across the glycaemic/BMI strata. The proportions of individuals achieving LDL cholesterol thresholds of <1.8 mmol/L and <1.4 mmol/L with inclisiran increased with increasing glycaemic and BMI strata. Across the glycaemic/BMI strata, a higher proportion of individuals had mild/moderate treatment-emergent adverse events (TEAEs) at the injection site with inclisiran (2.8%-7.7%) versus placebo (0.2%-2.1%)., Conclusion: Inclisiran provided substantial and sustained LDL cholesterol lowering across glycaemic/BMI strata, with a modest excess of transient mild-to-moderate TEAEs at the injection site., (© 2024 The Authors. Diabetes, Obesity and Metabolism published by John Wiley & Sons Ltd.)

The prognosis after heart transplantation continues to improve. Therefore, the prevention of chronic post-transplant sequelae, such as chronic kidney disease, allograft vasculopathy, and malignancies is becoming increasingly important. Everolimus, an inhibitor of the mammalian target of rapamycin (mTOR), is increasingly used for immunosuppression after heart transplantation. However, everolimus may cause a characteristic complex of adverse effects, including dyslipidemia. Currently there are no guidelines for the long-term screening and treatment of dyslipidemia in heart transplant recipients treated with everolimus. This article presents a clinical case of hypercholesterolemia that developed after the start of the everolimus treatment in a heart recipient. The patient was a 39-year-old man who underwent orthotopic heart transplantation for ischemic cardiomyopathy in 2012 (at the age of 27). In 2019, the patient's immunosuppressive therapy was converted from mycophenolate mofetil to everolimus due to the development of cardiac allograft vasculopathy. The change in the immunosuppressive therapy was associated with increases in total cholesterol and low-density lipoprotein cholesterol, which were not reversed with a combined lipid-lowering therapy (maximum doses of rosuvastatin, ezetimibe, fenofibrate). A decrease in lipid levels was achieved with a blocker of hepatic proprotein convertase subtilisin/kexin type 9 synthesis at the level of microribonucleic acid (inclisiran). This case demonstrates the difficulties in correcting dyslipidemia in patients with cardiac allograft, since the treatment with the immunosuppressant everolimus worsens existing dyslipidemia. However, the combination lipid-lowering therapy, that affects various elements of the pathogenesis (specifically, the combined inhibition of hydroxymethylglutaryl-CoA reductase with a statin, cholesterol absorption from the small intestine with ezetimibe, and PCSK9 messenger RNA with inclisiran), provides an effective control of blood lipids and minimizing the adverse effects of immunosuppressive therapy, such as cardiac allograft vasculopathy.

Bacterial diseases are one of the most common issues that result in crop loss worldwide, and the increasing usage of chemical pesticides has caused the occurrence of resistance in pathogenic bacteria and environmental pollution problems. Nanomaterial mediated gene silencing is starting to display powerful efficiency and environmental friendliness for improving plant disease resistance. However, the internalization of nanomaterials and the physiological mechanisms behind nano-improved plant disease resistance are still rarely understood. We engineered the polyethyleneimine (PEI) functionalized gold nanoparticles (PEI-AuNPs) with fluorescent properties and ROS scavenging activity to act as siRNA delivery platforms. Besides the loading, protection, and delivery of nucleic acid molecules in plant mature leaf cells by PEI-AuNPs, its fluorescent property further enables the traceability of the distribution of the loaded nucleic acid molecules in cells. Additionally, the PEI-AuNPs-based RNAi delivery system successfully mediated the silencing of defense-regulated gene AtWRKY1. Compared to control plants, the silenced plants performed better resistance to Pseudomonas syringae, showing a reduced bacterial number, decreased ROS content, increased antioxidant enzyme activities, and improved chlorophyll fluorescence performance. Our results showed the advantages of AuNP-based RNAi technology in improving plant disease resistance, as well as the potential of plant nanobiotechnology to protect agricultural production., (© 2024. The Author(s).)

Background: MutT homolog 1 (MTH1) sanitizes oxidized dNTP pools to promote the survival of cancer cells and its expression is frequently upregulated in cancers. Polyubiquitination stabilizes MTH1 to facilitate the proliferation of melanoma cells, suggesting the ubiquitin system controls the stability and function of MTH1. However, whether ubiquitination regulates MTH1 in gastric cancers has not been well defined. This study aims to investigate the interaction between MTH1 and a deubiquitinase, USP9X, in regulating the proliferation, survival, migration, and invasion of gastric cancer cells., Methods: The interaction between USP9X and MTH1 was evaluated by co-immunoprecipitation (co-IP) in HGC-27 gastric cancer cells. siRNAs were used to interfere with USP9X expression in gastric cancer cell lines HGC-27 and MKN-45. MTT assays were carried out to examine the proliferation, propidium iodide (PI) and 7-AAD staining assays were performed to assess the cell cycle, Annexin V/PI staining assays were conducted to examine the apoptosis, and transwell assays were used to determine the migration and invasion of control, USP9X-deficient, and USP9X-deficient plus MTH1-overexpressing HGC-27 and MKN-45 gastric cancer cells., Results: Co-IP data show that USP9X interacts with and deubiquitinates MTH1. Overexpression of USP9X elevates MTH1 protein level by downregulating its ubiquitination, while knockdown of USP9X has the opposite effect on MTH1. USP9X deficiency in HGC-27 and MKN-45 cells causes decreased proliferation, cell cycle arrest, extra apoptosis, and defective migration and invasion, which could be rescued by excessive MTH1., Conclusion: USP9X interacts with and stabilizes MTH1 to promote the proliferation, survival, migration and invasion of gastric cancer cells., (© 2024. The Author(s).)

Pulmonary Fibrosis (PF) is a fatal disease in the interstitial lung associated with high mortality, morbidity, and poor prognosis. Transforming growth factor-β1 (TGF-β1) is a fibroblast-activating protein that promotes fibrous diseases. Herein, an inhalable system was first developed using milk exosomes (M-Exos) encapsulating siRNA against TGF-β1 (MsiTGF-β1), and their therapeutic potential for bleomycin (BLM)-induced PF was investigated. M-siTGF-β1 was introduced into the lungs of mice with PF through nebulization. The collagen penetration effect and lysosomal escape ability were verified in vitro. Inhaled MsiTGF-β1 notably alleviated inflammatory infiltration, attenuated extracellular matrix (ECM) deposition, and increased the survival rate of PF mice by 4.7-fold. M-siTGF-β1 protected lung tissue from BLM toxicity by efficiently delivering specific siRNA to the lungs, leading to TGF-β1 mRNA silencing and epithelial mesenchymal transition pathway inhibition. Therefore, M-siTGF-β1 offers a promising avenue for therapeutic intervention in fibrosis-related disorders., (© 2024. The Author(s).)

Background and Objectives: Lowering low-density lipoprotein (LDL-C) levels is critical for preventing atherosclerotic cardiovascular disease, yet some patients fail to reach the LDL-C targets despite available intensive lipid-lowering therapies. This study assessed the effectiveness and safety profile of alirocumab, evolocumab, and inclisiran in lipid reduction. Materials and Methods: A cohort of 51 patients (median (Q1-Q3) age: 49.0 (39.5-57.5) years) was analyzed. Eligibility included an LDL-C level > 2.5 mmol/L while on the maximum tolerated dose of statin and ezetimibe, a diagnosis of familial hypercholesterolemia, or a very high risk of cardiovascular diseases following myocardial infarction within 12 months prior to the study. Follow-ups and lab assessments were conducted at baseline (51 patients), 3 months (51 patients), and 15 months (26 patients) after the treatment initiation. Results: Median initial LDL-C levels 4.1 (2.9-5.0) mmol/L, decreasing significantly to 1.1 (0.9-1.6) mmol/L at 3 months and 1.0 (0.7-1.8) mmol/L at 15 months ( p < 0.001). Total cholesterol also reduced significantly compared to baseline at both intervals ( p < 0.001). No substantial differences in LDL-C or total cholesterol levels were observed between 3- and 15-month observations ( p > 0.05). No statistically significant differences were noted in cholesterol reduction among the alirocumab, evolocumab, and inclisiran groups at 3 months. The safety profile was favorable, with no reported adverse cardiovascular events or significant changes in alanine transaminase, creatinine, or creatine kinase levels. Conclusions: Alirocumab, evolocumab, and inclisiran notably decreased LDL-C and total cholesterol levels without significant adverse effects, underscoring their potential as effective treatments in patients who do not achieve lipid targets with conventional therapies.

The fracture healing outcome is largely dependent on the quantities as well as osteogenic differentiation capacities of mesenchymal stem cells (MSCs) at the lesion site. Herein, macrophage membrane (MM)-reversibly cloaked nanocomplexes (NCs) are engineered for the lesion-targeted and hierarchical co-delivery of short stromal derived factor-1α peptide (sSDF-1α) and Ckip-1 small interfering RNA (Ckip-1 siRNA, siCkip-1) to promote bone repair by concurrently fostering recruitment and osteogenic differentiation of endogenous MSCs. To construct the NCs, a membrane-penetrating α-helical polypeptide first assembles with siCkip-1, and the cationic NCs are sequentially coated with catalase and an outer shell of sSDF-1α-anchored MM. Due to MM-assisted inflammation homing, intravenously injected NCs could efficiently accumulate at the fractured femur, where catalase decomposes the local hydrogen peroxide to generate oxygen bubbles that drives the shedding of sSDF-1α-anchored MM in the extracellular compartment. The exposed, cationic inner core thus enables robust trans-membrane delivery into MSCs to induce Ckip-1 silencing. Consequently, sSDF-1α-guided MSCs recruitment cooperates with siCkip-1-mediated osteogenic differentiation to facilitate bone formation and accelerate bone fracture healing. This study provides an enlightened strategy for the hierarchical co-delivery of macromolecular drugs into different cellular compartments, and it also renders a promising modality for the management of fracture healing., (© 2024. The Author(s).)

Objective: This study aimed to investigate the critical role of MDSCs in CRC immune suppression, focusing on the CSF1R and JAK/STAT3 signaling axis. Additionally, it assessed the therapeutic efficacy of LNCs@CSF1R siRNA and anti-PD-1 in combination., Methods: Single-cell transcriptome sequencing data from CRC and adjacent normal tissues identified MDSC-related differentially expressed genes. RNA-seq analysis comprehensively profiled MDSC gene expression in murine CRC tumors. LNCs@CSF1R siRNA nanocarriers effectively targeted and inhibited CSF1R. Flow cytometry quantified changes in MDSC surface markers post-CSF1R inhibition. RNA-seq and pathway enrichment analyses revealed the impact of CSF1R on MDSC metabolism and signaling. The effect of CSF1R inhibition on the JAK/STAT3 signaling axis was validated using Colivelin and metabolic assessments. Glucose and fatty acid uptake were measured via fluorescence-based flow cytometry. The efficacy of LNCs@CSF1R siRNA and anti-PD-1, alone and in combination, was evaluated in a murine CRC model with extensive tumor section analyses., Results: CSF1R played a significant role in MDSC-mediated immune suppression. LNCs@CSF1R siRNA nanocarriers effectively targeted MDSCs and inhibited CSF1R. CSF1R regulated MDSC fatty acid metabolism and immune suppression through the JAK/STAT3 signaling axis. Inhibition of CSF1R reduced STAT3 activation and target gene expression, which was rescued by Colivelin. Combined treatment with LNCs@CSF1R siRNA and anti-PD-1 significantly slowed tumor growth and reduced MDSC abundance within CRC tumors., Conclusion: CSF1R via the JAK/STAT3 axis critically regulates MDSCs, particularly in fatty acid metabolism and immune suppression. Combined therapy with LNCs@CSF1R siRNA and anti-PD-1 enhances therapeutic efficacy in a murine CRC model, providing a strong foundation for future clinical applications., (© 2024. The Author(s).)

Macrophages show high plasticity and play a vital role in the progression of metabolic dysfunction-associated steatohepatitis (MASH). X-box binding protein 1 (XBP1), a key sensor of the unfolded protein response, can modulate macrophage-mediated pro-inflammatory responses in the pathogenesis of MASH. However, how XBP1 influences macrophage plasticity and promotes MASH progression remains unclear. Herein, we formulated an Xbp1 siRNA delivery system based on folic acid modified D-α-tocopheryl polyethylene glycol 1000 succinate nanoparticles (FT@XBP1) to explore the precise role of macrophage-specific Xbp1 deficiency in the progression of MASH. FT@XBP1 was specifically internalized into hepatic macrophages and subsequently inhibited the expression of spliced XBP1 both in vitro and in vivo . It promoted M1-phenotype macrophage repolarization to M2 macrophages, reduced the release of pro-inflammatory factors, and alleviated hepatic steatosis, liver injury, and fibrosis in mice with fat-, fructose- and cholesterol-rich diet-induced MASH. Mechanistically, FT@XBP1 promoted macrophage polarization toward the M2 phenotype and enhanced the release of exosomes that could inhibit the activation of hepatic stellate cells. A promising macrophage-targeted siRNA delivery system was revealed to pave a promising strategy in the treatment of MASH., Competing Interests: Competing Interests: The authors have declared that no competing interest exists., (© The author(s).)

Pancreatic ductal adenocarcinoma (PDAC) is characterised by immune evasion that contribute to poor prognosis. Cancer-associated fibroblasts (CAFs) play a pivotal role in orchestrating the PDAC tumour microenvironment. We investigated the role of CAF-derived extracellular vesicle (EV)-packaged long non-coding RNAs (lncRNAs) in immune evasion and explored gene therapy using engineered EVs loading small interfering RNAs (siRNAs) as a potential therapeutic strategy. Our findings highlight the significance of EV-packaged lncRNA RP11-161H23.5 from CAF in promoting PDAC immune evasion by downregulating HLA-A expression, a key component of antigen presentation. Mechanistically, RP11-161H23.5 forms a complex with CNOT4, a subunit of the mRNA deadenylase CCR4-NOT complex, enhancing the degradation of HLA-A mRNA by shortening its poly(A) tail. This immune evasion mechanism compromises the anti-tumour immune response. To combat this, we propose an innovative approach utilising engineered EVs as natural and biocompatible nanocarriers for siRNA-based gene therapy and this strategy holds promise for enhancing the effectiveness of immunotherapy in PDAC. Overall, our study sheds light on the critical role of CAF-derived EV-packaged lncRNA RP11-161H23.5/CNOT4/HLA-A axis in PDAC immune evasion and presents a novel avenue for therapeutic intervention., (© 2024 The Author(s). Journal of Extracellular Vesicles published by Wiley Periodicals LLC on behalf of International Society for Extracellular Vesicles.)

With the emergence of gene therapy utilizing viral vectors, the potential risks associated with these vectors have prompted increased attention toward non-viral alternatives. DNA nanotechnology enables the assembly of specific oligonucleotide chains into nanostructures possessing defined spatial configurations. Due to their inherent characteristics, DNA nanostructures possess natural advantages as carriers for regulating gene expression in a non-viral manner. Cholesterol modification can convert DNA nanostructures from hydrophilic materials to amphiphilic materials, thereby extending their systemic circulation time. In this study, the high-dimensional design and cholesterol modification are shown to prolong the systemic circulation half-life of DNA nanostructures in mice. Specifically, the tetrahedron structure modified with three cholesterol molecules (TDN-3Chol) exhibit excellent circulation time and demonstrate a preference for renal uptake. The unique characteristics of TDN-3Chol can effectively deliver p53 siRNA to the mouse renal tubular tissue, resulting in successful knockdown of p53 and demonstrating its potential for preventing acute kidney injury. Furthermore, TDN-3Chol is not exhibited significant toxicity in mice, highlighting its promising role as a non-viral vector for targeted gene expression regulation in the kidneys. The designed non-viral vector as a prophylactic medication shows potential in addressing the current clinical challenges associated with nephrotoxic drugs., (© 2024 Wiley‐VCH GmbH.)

Diagnosis and treatment of tumor especially drug-resistant tumor remains a huge challenge, which requires intelligent nanomedicines with low toxic side effects and high efficacy. Herein, deformable smart DNA nanomachines are developed for synergistic intracellular cancer-related miRNAs imaging and chemo-gene therapy of drug-resistant tumors. The tetrahedral DNA framework (MA-TDNA) with fluorescence quenched component and five antennas is self-assembled first, and then DOX molecules are loaded on the MA-TDNAs followed by linking MUC1-aptamer and Mcl-1 siRNA to the antennas of MA-TDNA, so that the apt-MA-TDNA@DOX-siRNA (DNA nanomachines) is constructed. The DNA nanomachine can respond to two tumor-related miRNAs in vitro and in vivo, which can undergo intelligent miRNA-triggered opening of the framework, resulting in the "turn on" of the fluorescence for sensitively and specifically sensing intracellular miRNAs. Meanwhile, both miRNA-responded rapid release and pH-responded release of DOX are achieved for chemotherapy of tumor. In addition, the gene therapy of the DNA nanomachines is achieved due to the miRNA-specific capture and the RNase H triggered release of Mcl-1 siRNA. The DNA nanomachines intergrading both tumor imaging and chemo-gene therapy in single nanostructures realized efficient tumor-targeted, image-guided, and microenvironment-responsive tumor diagnosis and treatment, which provides a synergetic antitumor effect on drug-resistant tumor., (© 2024 Wiley‐VCH GmbH.)

The successful translation of therapeutic nucleic acids (NAs) for the treatment of neurological disorders depends on their safe and efficient delivery to neural cells, in particular neurons. DNA nanostructures can be a promising NAs delivery vehicle. Nonetheless, the potential of DNA nanostructures for neuronal cell delivery of therapeutic NAs is unexplored. Here, tetrahedral DNA nanostructures (TDN) as siRNA delivery scaffolds to neuronal cells, exploring the influence of functionalization with two different reported neuronal targeting ligands: C4-3 RNA aptamer and Tet1 peptide are investigated. Nanostructures are characterized in vitro, as well as in silico using molecular dynamic simulations to better understand the overall TDN structural stability. Enhancement of neuronal cell uptake of TDN functionalized with the C4-3 Aptamer (TDN-Apt), not only in neuronal cell lines but also in primary neuronal cell cultures is demonstrated. Additionally, TDN and TDN-Apt nanostructures carrying siRNA are shown to promote silencing in a process aided by chloroquine-induced endosomal disruption. This work presents a thorough workflow for the structural and functional characterization of the proposed TDN as a nano-scaffold for neuronal delivery of therapeutic NAs and for targeting ligands evaluation, contributing to the future development of new neuronal drug delivery systems based on DNA nanostructures., (© 2024 The Authors. Small published by Wiley‐VCH GmbH.)

Competing Interests: Declaration of Competing Interest None.

Cardiovascular disease (CVD) is the leading cause of morbidity and mortality worldwide. Hyperlipidemia, particularly elevated low-density lipoprotein cholesterol (LDL-C) is one of the major risk factors for CVD. Major landmark cardiovascular outcome clinical trials demonstrated that LDL-C lowering medications reduce cardiovascular events, and the lower the LDL-C the better the outcome. This article discusses the evolution of LDL-C lowering medications starting from bile acid sequestrants (BAS), statin therapy, bempedoic acid, the proprotein convertase subtilisin kexin 9 (PCSK9) synthesis inhibitor, novel small interfering RNA-based therapy (inclisiran) to the most recent oral PCSK9 inhibitors (MK-0616) which is currently under phase 3 clinical trial studies., Competing Interests: Declaration of competing interest All authors declare no conflict of interest. This manuscript is not under consideration elsewhere., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Low-expression antigens, especially neoantigens, pose a significant challenge in immunotherapy for low immunogenicity pancreatic cancer. Increasing the tumor mutation burden is crucial to enhance the expression of tumor antigens and improve tumor immunogenicity. However, the incomplete intervention in DNA stability hampers effective elevation of the tumor mutation burden, thus reducing the probability of neoantigen. To address this issue, we have developed a novel nano-regulator that intervenes in the DNA stability of tumor cells, thereby enhancing tumor mutations. This nano-regulator comprises metal-organic frameworks (MOFs) encapsulating DNA damage agent doxorubicin and DNA damage repair inhibitor siRNA-ATR, enabling simultaneous induction of DNA mutations and inhibition of their repair. Importantly, this regulator, named as MOF DOX&siATR , can modulate the tumor gene expression profile, induce the production of neoantigens of Atp8b1, and enhance the immunogenicity of pancreatic cancer. The characteristics of DNA stability intervention by MOF DOX&siATR hold promise for augmenting the immune response in low immunogenic tumors, making it a potential nanomedicine for the treatment of pancreatic cancer., 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 Ltd. All rights reserved.)

Objective: To investigate the effects of cigarette smoke (CS) on Serine/Threonine Kinase 11 (STK11) and to determine STK11's role in CS-induced airway epithelial cell cytotoxicity. Methods: STK11 expression levels in the lung tissues of smokers with or without COPD and mice exposed to CS or room air (RA) were determined by immunoblotting and RT-PCR. BEAS-2Bs-human bronchial airway epithelial cells were exposed to CS extract (CSE), and the changes in STK11 expression levels were determined by immunoblotting and RT-PCR. BEAS-2B cells were transfected with STK11-specific siRNA or STK11 expression plasmid, and the effects of CSE on airway epithelial cell cytotoxicity were measured. To determine the specific STK11 degradation-proteolytic pathway, BEAS-2Bs were treated with cycloheximide alone or combined with MG132 or leupeptin. Finally, to identify the F-box protein mediating the STK11 degradation, a screening assay was performed using transfection with a panel of FBXL E3 ligase subunits. Results: STK11 protein levels were significantly decreased in the lung tissues of smokers with COPD relative to smokers without COPD. STK11 protein levels were also significantly decreased in mouse lung tissues exposed to CS compared to RA. Exposure to CSE shortened the STK11 mRNA and protein half-life to 4 h in BEAS-2B cells. STK11 protein overexpression attenuated the CSE-induced cytotoxicity; in contrast, its knockdown augmented CSE-induced cytotoxicity. FBXL19 mediates CSE-induced STK11 protein degradation via the ubiquitin-proteasome pathway in cultured BEAS-2B cells. FBXL19 overexpression led to accelerated STK11 ubiquitination and degradation in a dose-dependent manner. Conclusions: Our results suggest that CSE enhances the degradation of STK11 protein in airway epithelial cells via the FBXL19-mediated ubiquitin-proteasomal pathway, leading to augmented cell death.HIGHLIGHTSLung tissues of COPD-smokers exhibited a decreased STK11 RNA and protein expression.STK11 overexpression attenuates CS-induced airway epithelial cell cytotoxicity.STK11 depletion augments CS-induced airway epithelial cell cytotoxicity.CS diminishes STK11 via FBXL19-mediated ubiquitin-proteasome degradation.

Background: More than 80% of patients may experience acute pain after a surgical procedure, and this is often refractory to pharmacological intervention. The identification of new targets to treat postoperative pain is necessary. There is an association of polymorphisms in the Ca v 2.3 gene with postoperative pain and opioid consumption. Our study aimed to identify Ca v 2.3 as a potential target to treat postoperative pain and to reduce opioid-related side effects., Experimental Approach: A plantar incision model was established in adult male and female C57BL/6 mice. Ca v 2.3 expression was detected by qPCR and suppressed by siRNA treatment. The antinociceptive efficacy and safety of a Ca v 2.3 blocker-alone or together with morphine-was also assessed after surgery., Key Results: Paw incision in female and male mice caused acute nociception and increased Ca v 2.3 mRNA expression in the spinal cord but not in the incised tissue. Intrathecal treatment with siRNA against Ca v 2.3, but not with a scrambled siRNA, prevented the development of surgery-induced nociception in both male and female mice, with female mice experiencing long-lasting effects. High doses of i.t. SNX-482, a Ca v 2.3 channel blocker, or morphine injected alone, reversed postoperative nociception but also induced side effects. A combination of lower doses of morphine and SNX-482 mediated a long-lasting reversal of postsurgical pain in female and male mice., Conclusion: Our results demonstrate that Ca v 2.3 has a pronociceptive role in the induction of postoperative pain, indicating that it is a potential target for the development of therapeutic approaches for the treatment of postoperative pain., (© 2024 British Pharmacological Society.)

Tumor-associated macrophages (TAMs) are a promising target for cancer immunotherapy, but delivering therapeutic agents to TAMs within the tumor microenvironment (TME) is challenging. In this study, a photosensitive, dual-targeting nanoparticle system (M.RGD@Cr-CTS-siYTHDF1 NPs) was developed. The structure includes a shell of DSPE-modified RGD peptides targeting integrin receptors on tumor cells and carboxymethyl mannose targeting CD206 receptors on macrophages, with a core of chitosan adsorbing m6A reading protein YTHDF1 siRNA and chromium nanoparticles (Cr NPs). The approach is specifically designed to target TAM and cancer cells, utilizing the photothermal effect of Cr NPs to disrupt the TME and deliver siYTHDF1 to TAM. In experiments with tumor-bearing mice, M.RGD@Cr-CTS-siYTHDF1 NPs, when exposed to laser irradiation, effectively killed tumor cells, disrupted the TME, delivered siYTHDF1 to TAMs, silenced the YTHDF1 gene, and shifted the STAT3-STAT1 equilibrium by reducing STAT3 and enhancing STAT1 expression. This reprogramming of TAMs towards an anti-tumor phenotype led to a pro-immunogenic TME state. The strategy also suppressed immunosuppressive IL-10 production, increased expression of immunostimulatory factors (IL-12 and IFN-γ), boosted CD8 + T cell infiltration and M1-type TAMs, and reduced Tregs and M2-type TAMs within the TME. In conclusion, the dual-targeting M.RGD@Cr-CTS-siYTHDF1 NPs, integrating dual-targeting capabilities with photothermal therapy (PTT) and RNA interference, offer a promising approach for molecular targeted cancer immunotherapy with potential for clinical application., (© 2024. The Author(s).)

Background and Objectives : This study aimed to investigate the relationship between neuropathic pain and CREB-binding protein (CBP) and methyl-CpG-binding protein 2 (MeCP2) expression levels in a rat model with spared nerve injury (SNI). Materials and Methods : Rat (male Sprague-Dawley white rats) models with surgical SNI (n = 6) were prepared, and naive rats (n = 5) were used as controls. The expression levels of CBP and MeCP2 in the spinal cord and dorsal root ganglion (DRG) were compared through immunohistochemistry at 7 and 14 days after surgery. The relationship between neuropathic pain and CBP/MeCP2 was also analyzed through intrathecal siRNA administration. Results : SNI induced a significant increase in the number of CBPs in L4 compared with contralateral DRG as well as with naive rats. The number of MeCP2 cells in the dorsal horn on the ipsilateral side decreased significantly compared with the contralateral dorsal horn and the control group. SNI induced a significant decrease in the number of MeCP2 neurons in the L4 ipsilateral DRG compared with the contralateral DRG and naive rats. The intrathecal injection of CBP siRNA significantly inhibited mechanical allodynia induced by SNI compared with non-targeting siRNA treatment. MeCP2 siRNA injection showed no significant effect on mechanical allodynia. Conclusions : The results suggest that CBP and MeCP2 may play an important role in the generation of neuropathic pain following peripheral nerve injury.

Background: Plasma concentration of PAI-1 (plasminogen activator inhibitor-1) correlates with arterial stiffness. Vascular smooth muscle cells (SMCs) express PAI-1, and the intrinsic stiffness of SMCs is a major determinant of total arterial stiffness. We hypothesized that PAI-1 promotes SMC stiffness by regulating the cytoskeleton and that pharmacological inhibition of PAI-1 decreases SMC and aortic stiffness., Methods: PAI-039, a specific inhibitor of PAI-1, and small interfering RNA were used to inhibit PAI-1 expression in cultured human SMCs. Effects of PAI-1 inhibition on SMC stiffness, F-actin (filamentous actin) content, and cytoskeleton-modulating enzymes were assessed. WT (wild-type) and PAI-1-deficient murine SMCs were used to determine PAI-039 specificity. RNA sequencing was performed to determine the effects of PAI-039 on SMC gene expression. In vivo effects of PAI-039 were assessed by aortic pulse wave velocity., Results: PAI-039 significantly reduced intrinsic stiffness of human SMCs, which was accompanied by a significant decrease in cytoplasmic F-actin content. PAI-1 gene knockdown also decreased cytoplasmic F-actin. PAI-1 inhibition significantly increased the activity of cofilin, an F-actin depolymerase, in WT murine SMCs, but not in PAI-1-deficient SMCs. RNA-sequencing analysis suggested that PAI-039 upregulates AMPK (AMP-activated protein kinase) signaling in SMCs, which was confirmed by Western blotting. Inhibition of AMPK prevented activation of cofilin by PAI-039. In mice, PAI-039 significantly decreased aortic stiffness and tunica media F-actin content without altering the elastin or collagen content., Conclusions: PAI-039 decreases intrinsic SMC stiffness and cytoplasmic stress fiber content. These effects are mediated by AMPK-dependent activation of cofilin. PAI-039 also decreases aortic stiffness in vivo. These findings suggest that PAI-1 is an important regulator of the SMC cytoskeleton and that pharmacological inhibition of PAI-1 has the potential to prevent and treat cardiovascular diseases involving arterial stiffening.

Skin wound healing is driven by proliferation, migration and differentiation of several cell types that are controlled by the alterations in the gene expression programmes. Brahma Gene 1 (BRG1) (also known as SMARCA4) is a core ATPase in the BRG1 Associated Factors (BAF) ATP-dependent chromatin remodelling complexes that alter DNA-histone interaction in chromatin at the specific gene regulatory elements resulting in increase or decrease of the target gene transcription. Using siRNA mediated suppression of BRG1 during wound healing in a human ex vivo and in vitro (scratch assay) models, we demonstrated that BRG1 is essential for efficient skin wound healing by promoting epidermal keratinocytes migration, but not their proliferation or survival. BRG1 controls changes in the expression of genes associated with gene transcription, response to wounding, cell migration and cell signalling. Altogether, our data revealed that BRG1 play positive role in skin repair by promoting keratinocyte migration and impacting the genes expression programmes associated with cell migration and cellular signalling., (© 2024 Unilever U.K. Central Resources Limited and The Author(s). Experimental Dermatology published by John Wiley & Sons Ltd.)