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Endoglin (ENG) mutation causes type 1 hereditary hemorrhagic telangiectasia (HHT1). HHT1 patients have arteriovenous malformations (AVMs) in multiple organs, including the brain. In mice, Eng deletion induced by R26RCreER or SM22αCre leads to AVM development in the brain and other organs. We hypothesized that an increase in Eng- negative ECs will enhance AVM severity. To increase EC Eng deletion, we used a codon-improved cre (icre), which is more potent in recombination of the floxed alleles than the wild-type (WT) cre. R26RCreER;Engf/f mice that have a Rosa promoter driving and tamoxifen (TM)-inducible WT cre expression globally, and PdgfbiCreER;Engf/f mice that have a Pdgfb promoter driving and TM-inducible icre expression in ECs were treated with three intra-peritoneal injections of TM (2.5 mg/25 g of body weight) to delete Eng globally or in the ECs. AAV-VEGF was stereotactically injected into the brain to induce brain focal angiogenesis and brain AVM. We found that icre caused more Eng deletion in the brain, indicated by a lower level of Eng proteins (p < 0.001) and fewer Eng-positive ECs (p = 0.01) than mice with WT cre. Mice with icre-mediated Eng deletion have more abnormal vessels (p = 0.02), CD68+ macrophages (p = 0.002), and hemorrhage (p = 0.04) and less vascular pericyte and smooth muscle coverage than mice with WT cre. In addition, arteriovenous shunts were detected in the intestines of icre mice, a phenotype that has not been detected in WT cre mice before. RNA-seq analysis showed that 8 out of the 10 top upregulated pathways identified by gene ontology (GO) analysis are related to inflammation. Therefore, the increase in Eng deletion in ECs exacerbates AVM severity, which is associated with enhanced inflammation. Strategies that can reduce Eng-negative ECs could be used to develop new therapies to reduce AVM severity for HHT1 patients.

Background: Hereditary Hemorrhagic Telangiectasia (HHT) is a genetic disorder leading to frequent bleeding in several organs. As HHT diagnosis is demanding and depends on clinical criteria, liquid biopsy would be beneficial. Exosomes from biofluids are nano-sized vesicles for intercellular communication. Their cargo and characteristics represent biomarkers for many diseases. Here, exosomes of HHT patients were examined regarding their biosignature. Methods: Exosomes were isolated from the plasma of 20 HHT patients and 17 healthy donors (HDs). The total exosomal protein was quantified, and specific proteins were analyzed using Western blot and antibody arrays. Human umbilical vein endothelial cells (HUVECs) co-incubated with exosomes were functionally examined via immunofluorescence, proliferation, and scratch assay. Results: The levels of the angiogenesis-regulating protein Thrombospondin-1 were significantly higher in HHT compared to HD exosomes. Among HHT, but not HD exosomes, a negative correlation between total exosomal protein and soluble Endoglin (sENG) levels was found. Other exosomal proteins (ALK1, ALK5) and the particle concentration significantly correlated with disease severity parameters (total consultations/interventions, epistaxis severity score) in HHT patients. Functionally, HUVECs were able to internalize both HD and HHT exosomes, inducing a similar change in the F-Actin structure and a reduction in migration and proliferation. Conclusions: This study provided first insights into the protein cargo and function of HHT-derived exosomes. The data indicate changes in sENG secretion via exosomes and reveal exosomal Thrombospondin-1 as a potential biomarker for HHT. Several exosomal characteristics were pointed out as potential liquid biomarkers for disease severity, revealing a possible new way of diagnosis and prognosis of HHT. [ABSTRACT FROM AUTHOR]

Objectives: This study aims to investigate the expression and role of Endoglin (ENG) in endometriosis (EM). Material and methods: In this study, quantitative real-time polymerase chain reaction, western blot and immunohistochemistry were used to examine the expression of ENG in tissues. Cellular experiments were performed to evaluate the effect of ENG on cellular biological function. Western blot was used to examine the expression of epithelial to mesenchymal transition-related proteins. Results: The expression of ENG was significantly higher in the ectopic endometriotic tissues than that in eutopic endometriotic tissues. Knockdown of ENG inhibited cell viability, migration and invasion, and induced cell apoptosis in hEM15A cells. Additionally, silenced ENG caused increased levels of E-cadherin and decreased levels of N-cadherin, vimentin, MMP-2 and MMP-9. Conclusions: These results confirmed that ENG may be involved in the development of endometriosis by promoting EMT process, revealing a new insight into the pathogenesis of endometriosis and contributing to the exploration of molecular therapeutic strategies against endometriosis. [ABSTRACT FROM AUTHOR]

Background: Angiogenic factor imbalance, such as that caused by soluble endoglin (sEng), is a feature of preeclampsia. However, the connection between sEng and clinical and laboratory indicators, and the severity of preeclampsia is not entirely understood. Methods: Ninety subjects were incorporated in this study, 30 were healthy pregnant with mean age (35.6) years. (32) mild Preeclampsia case with mean age (32.65) years preeclampsia, and (30) sever PE with mean age (32.65) wherever all women were more than 24 weeks of pregnancy. Mean blood pressure and level of proteinuria were used as indicators of the severity of the disease. ELISA was used to measure the levels of sEng in the serum. Serum lipid profile was measured by enzymatic methods. The qualitative dip-stick technique (CYBOWTM DFI Co Ltd, Republic of Korea) was used to assess the amount of urine protein. Results: A substantial difference between the PE group's serum sEng concentration and that of the healthy subjects (p 0.001) could be seen. Serum sEng concentrations of patients with moderate PE and those with severe PE differed significantly. Additionally, there were strong positive relationships between the serum sEng concentration and the SBP and proteinuria. serum sEng levels, biochemical indicators, and other factors, however, did not significantly correlate. The diagnostic accuracy (86.2) in distinguishing mild PE from the healthy patient group was good thanks to the excellent area under the curve (AUC = 0.827, p < 0.0001). Conclusion: Levels of sEng in sera of PE patients were elevated. sEng was significantly correlated with PE patients. [ABSTRACT FROM AUTHOR]

Abstract Background In vitro disease modeling enables translational research by providing insight into disease pathophysiology and molecular mechanisms, leading to the development of novel therapeutics. Nevertheless, in vitro systems have limitations for recapitulating the complexity of tissues, and a single model system is insufficient to gain a comprehensive understanding of a disease. Results Here we explored the potential of using several models in combination to provide mechanistic insight into hereditary hemorrhagic telangiectasia (HHT), a genetic vascular disorder. Genome editing was performed to establish hPSCs (H9) with ENG haploinsufficiency and several in vitro models were used to recapitulate the functional aspects of the cells that constitute blood vessels. In a 2D culture system, endothelial cells showed early senescence, reduced viability, and heightened susceptibility to apoptotic insults, and smooth muscle cells (SMCs) exhibited similar behavior to their wild-type counterparts. Features of HHT were evident in 3D blood-vessel organoid systems, including thickening of capillary structures, decreased interaction between ECs and surrounding SMCs, and reduced cell viability. Features of ENG haploinsufficiency were observed in arterial and venous EC subtypes, with arterial ECs showing significant impairments. Molecular biological approaches confirmed the significant downregulation of Notch signaling in HHT-ECs. Conclusions Overall, we demonstrated refined research strategies to enhance our comprehension of HHT, providing valuable insights for pathogenic analysis and the exploration of innovative therapeutic interventions. Additionally, these results underscore the importance of employing diverse in vitro systems to assess multiple aspects of disease, which is challenging using a single in vitro system.

Background and aimsDetailed investigation of the biological pathways leading to hepatic fibrosis and identification of liver fibrosis biomarkers may facilitate early interventions for pediatric cholestasis.Approach and resultsA targeted enzyme-linked immunosorbent assay-based panel of nine biomarkers (lysyl oxidase, tissue inhibitor matrix metalloproteinase (MMP) 1, connective tissue growth factor [CTGF], IL-8, endoglin, periostin, Mac-2-binding protein, MMP-3, and MMP-7) was examined in children with biliary atresia (BA; n = 187), alpha-1 antitrypsin deficiency (A1AT; n = 78), and Alagille syndrome (ALGS; n = 65) and correlated with liver stiffness (LSM) and biochemical measures of liver disease. Median age and LSM were 9 years and 9.5 kPa. After adjusting for covariates, there were positive correlations among LSM and endoglin ( p = 0.04) and IL-8 ( p

Background: Sepsis, a complex condition characterized by dysregulated immune response and organ dysfunction, is a leading cause of mortality in ICU patients. Current diagnostic and prognostic approaches primarily rely on non-specific biomarkers and illness severity scores, despite early endothelial activation being a key feature of sepsis. This study aimed to evaluate the levels of soluble thrombomodulin and soluble endoglin in seriously ill surgical septic patients and explore their association with organ dysfunction and disease severity. Methodology: A case control study was conducted from March 2022 to November 2022, involving seriously ill septic surgical patients. Baseline clinical and laboratory data were collected within 24 h of admission to the Surgical Intensive Care Unit. This included information such as age, sex, hemodynamic parameters, blood chemistry, SOFA score, qSOFA score, and APACHE-II score. A proforma was filled out to record these details. The outcome of each patient was noted at the time of discharge. Results: The study found significantly elevated levels of soluble thrombomodulin and soluble endoglin in seriously ill surgical septic patients. The RTqPCR analysis revealed a positive correlation between soluble thrombomodulin and soluble endoglin levels with the qSOFA score, as well as, there was a positive association between RTqPCR soluble thrombomodulin and the SOFA score. These findings indicate a correlation between these biomarkers and organ dysfunction and disease severity. Conclusion: The study concludes that elevated levels of soluble thrombomodulin and soluble endoglin can serve as endothelial biomarkers for early diagnosis and prognostication in seriously ill surgical septic patients. [ABSTRACT FROM AUTHOR]

Background: In vitro disease modeling enables translational research by providing insight into disease pathophysiology and molecular mechanisms, leading to the development of novel therapeutics. Nevertheless, in vitro systems have limitations for recapitulating the complexity of tissues, and a single model system is insufficient to gain a comprehensive understanding of a disease. Results: Here we explored the potential of using several models in combination to provide mechanistic insight into hereditary hemorrhagic telangiectasia (HHT), a genetic vascular disorder. Genome editing was performed to establish hPSCs (H9) with ENG haploinsufficiency and several in vitro models were used to recapitulate the functional aspects of the cells that constitute blood vessels. In a 2D culture system, endothelial cells showed early senescence, reduced viability, and heightened susceptibility to apoptotic insults, and smooth muscle cells (SMCs) exhibited similar behavior to their wild-type counterparts. Features of HHT were evident in 3D blood-vessel organoid systems, including thickening of capillary structures, decreased interaction between ECs and surrounding SMCs, and reduced cell viability. Features of ENG haploinsufficiency were observed in arterial and venous EC subtypes, with arterial ECs showing significant impairments. Molecular biological approaches confirmed the significant downregulation of Notch signaling in HHT-ECs. Conclusions: Overall, we demonstrated refined research strategies to enhance our comprehension of HHT, providing valuable insights for pathogenic analysis and the exploration of innovative therapeutic interventions. Additionally, these results underscore the importance of employing diverse in vitro systems to assess multiple aspects of disease, which is challenging using a single in vitro system. [ABSTRACT FROM AUTHOR]

Triple-negative breast cancer (TNBC) is characterized by the absence of the estrogen receptor, progesterone receptor, and receptor tyrosine kinase HER2 expression. Due to the limited number of FDA-approved targeted therapies for TNBC, there is an ongoing need to understand the molecular underpinnings of TNBC for the development of novel combinatorial treatment strategies. This study evaluated the role of the MerTK receptor tyrosine kinase on proliferation and invasion/metastatic potential in TNBC. Immunohistochemical analysis demonstrated MerTK expression in 58% of patient-derived TNBC xenografts. The stable overexpression of MerTK in human TNBC cell lines induced an increase in proliferation rates, robust in vivo tumor growth, heightened migration/invasion potential, and enhanced lung metastases. NanoString nCounter analysis of MerTK-overexpressing SUM102 cells (SUM102-MerTK) revealed upregulation of several signaling pathways, which ultimately drive cell cycle progression, reduce apoptosis, and enhance cell survival. Proteomic profiling indicated increased endoglin (ENG) production in SUM102-MerTK clones, suggesting that MerTK creates a conducive environment for increased proliferative and metastatic activity via elevated ENG expression. To determine ENG's role in increasing proliferation and/or metastatic potential, we knocked out ENG in a SUM102-MerTK clone with CRISPR technology. Although this ENG knockout clone exhibited similar in vivo growth to the parental SUM102-MerTK clone, lung metastasis numbers were significantly decreased ~4-fold, indicating that MerTK enhances invasion and metastasis through ENG. Our data suggest that MerTK regulates a unique proliferative signature in TNBC, promoting robust tumor growth and increased metastatic potential through ENG upregulation. Targeting MerTK and ENG simultaneously may provide a novel therapeutic approach for TNBC patients. [ABSTRACT FROM AUTHOR]

The interaction of natural killer and trophoblast cells underlies maternal-fetal immune tolerance. The data on the participation of endoglin (ENG, CD105), or its soluble form, in the regulation of the communication of these cells are currently insufficient. In this study, we have investigated the role of endoglin in the intercellular interactions between natural killer cells and trophoblasts. Here, we show that NK-92 cells and JEG-3 cells constitutively express MICA/B, and CD105. In the presence of JEG-3 cells, the expression of NKG2D, CD94, MICA/B, and CD105 by NK-92 cells was increased, and the number of NK-92 cells expressing NKG2A, CD94, and MICA was reduced. Antibodies against ENG and recombinant endoglin (rENG), attenuated the trophoblasts' influence and returned the phenotype of the NK-92 cells to that of cells found in monoculture conditions. The antibodies and rENG also increased the expression of pSMAD2/3 by NK cells in both monoculture and co-culture conditions. The antibodies increased the trophoblasts' sensitivity to the cytotoxic effect of NK cells. In general, our findings indicate a significant role of endoglin in the intercellular communication between NK cells and trophoblasts. We also speculate that endoglin forms a complex with TGFβ, which aids in TGFβ trafficking between these cells. [ABSTRACT FROM AUTHOR]

This study aimed to investigate maternal serum levels of some angiogenic factors and certain proteins in dairy cattle for (1) early prediction of unsuccessful fertilization and (2) early detection of possible pregnancy failures (early EM) after positive insemination Serum samples were collected from the same cattle at three distinct time points: 30 days before artificial insemination (B‐AI), on the day of artificial insemination (AI), and 30 days after artificial insemination (A‐AI). As a result of the pregnancy examination, the cows were divided into two main groups according to whether they were pregnant. The results showed that leucyl/cystinyl aminopeptidase (LNPEP) concentration was significantly decreased B‐AI and Secreted frizzled‐related proteins (SFRP‐3), Vascular Endothelial Growth Factor (VEGF) and LNPEP levels were significantly decreased on day of AI, while PRL level was increased, and these data have prognostic significance as early indicator of the risk of potentially failed pregnancy. Additionally, a significant decrease in LNPEP, SFRP3, and VEGF levels, along with an increase in PRL levels was also observed in A‐AI. These results suggest that these biomarkers can be used as a screening test to monitor the course of pregnancy. There were no significant differences in serum levels of Insulin‐Like Growth Factor 2 (IGF‐2), Tissue inhibitors of metalloproteinases (TIMP‐1), angiopoietin (ANG), Endoglin (ENG), Fibroblast growth factor (FGF), Inhibine‐A (INH‐A) and Transforming growth factors‐β1 (TGF‐β1) between the evaluated periods neither unsuccessful nor the successful pregnancy groups. This is the first study reporting that the maternal serum levels of LNPEP, SFRP3, VEGF, and PRL have important roles in pregnancy success and may indicate whether insemination outcome will be successful B‐AI and predict the risk of unsuccessful pregnancy after AI in dairy cattle. The increase in such studies will allow the development of more specific, practical, and applicable markers. [ABSTRACT FROM AUTHOR]

Sodium‐glucose cotransporter 2 inhibitors (SGLT2i) are rapidly gaining ground in the treatment of heart failure (HF) with reduced ejection fraction (HFrEF) and acute myocardial infarction (AMI) by an unknown mechanism. Upregulation of Na+/H+ exchanger 1 (NHE1), SGLT1, and Ca2+/calmodulin‐dependent protein kinase II (CaMKII) in the diseased hearts was found to be attenuated by prolonged SGLT2i treatment. Unfortunately, dapagliflozin is not well understood as to how Na+/Ca2+ homeostasis is affected in cardiomyocytes. In this study, we aimed to investigate whether mechanical stretch in cardiomyocytes upregulate SGLT2, resulted to loss of Na+/Ca2+ homeostasis via ERK and eNOS signaling. AMI (+) and AMI (−) serum levels were estimated using ELISA assays of TGFβ‐1 or endoglin (CD105). Human cardiomyocyte cell line AC16 was subjected to different stresses: 5% mild and 25% aggressive, at 1 Hz for 24 h. Immunofluorescence assays were used to estimate troponin I, CD105, SGLT1/2, eNOSS633, and ERK1/2T202/Y204 levels was performed for 5% (mild), and 25% elongation for 24 h. AMI (+) serum showed increased TGFβ1 and CD105 compared to AMI (−) patients. In consistent, troponin I, CD105, SGLT1/2, eNOSS633 and ERK1/2T202/Y204 were upregulated after 25% of 24 h cyclic stretch. Dapagliflozin addition caused SGLT2 inhibition, which significantly decreased troponin I, CD105, SGLT1/2, eNOSS633, and ERK1/2T202/Y204 under 25% cyclic stretching. In summary, SGLT2 may have sensed mechanical stretch in a way similar to cardiac overloading as in vivo. By blocking SGLT2 in stretched cardiomyocytes, the AMI biomarkers (CD105, troponin I and P‐ERK) were decreased, potentially to rescue eNOS production to maintain normal cellular function. This discovery of CD105 and SGLT2 increase in mechanically stretched cardiomyocytes suggests that SGLT2 may conceive a novel role in direct or indirect sensing of mechanical stretch, prompting the possibility of an in vitro cardiac overloaded cell model, an alternative to animal heart model. [ABSTRACT FROM AUTHOR]

Breast cancer has become the most common malignant tumor in women, and its diagnosis and treatment principles mainly depend on the traditional histological and molecular pathological features. However, the systemic metastasis of breast cancer begins when cancer cells invade the primary tumor site as well as surrounding blood and lymphatic vessels. Pathologists have been able to distinguish between lymphatic and vascular invasion through a combination of immunohistochemical and vascular endothelial labelling techniques, and have recognized the role of lymphatic endothelial cells and vascular endothelial cells in tumor progression and metastasis. This paper summarizes the relevant clinical research progress and application of specific lymphovascular endothelial markers in breast cancer in recent years, in order to provide ideas for further revealing the progress mechanism of breast cancer and exploring new prognostic indicators and therapeutic targets for breast cancer.

This study aimed to evaluate the clinicopathologic significance of the combined immunohistochemical expression of the epithelial-mesenchymal transition marker E-cadherin and the angiogenesis marker CD105 in laryngeal squamous cell carcinoma and assess correlation of their expression. Eighty-five patients who underwent complete resection as primary treatment were selected for this study. E-cadherin and CD105 expression levels were determined by immunohistochemistry. The receiver operating curve approach was applied to determine the cut-off value and separate patients with high and low expression of markers. The high-risk group (“CD105 high” and “E-cadherin low” expression) showed statistically significant correlations with age less than 66 years (p = 0.039), advanced T-status (T3–4) (p = 0.046), aggressive TNM stage (stage III–IV) (p = 0.003) and locoregional recurrence of disease (p = 0.004). In the Kaplan-Meier analyses, the high-risk group had significantly worse prognoses than other risk groups (log-rank test 2 = 9.415, p = 0.024). Spearman’s correlation coefficient analysis showed a nonsignificant negative correlation between the expression of E-cadherin and CD105 (rho = –0.073, p = 0.505). Simultaneous consideration of E-cadherin and CD105 is a simple panel of markers to determine aggressive tumour phenotype with a higher risk of disease recurrence in patients with laryngeal cancer.

BACKGROUND: The Eunice Kennedy Shriver National Institute of Child Health and Human Development Nulliparous Pregnancy Outcomes Study: Monitoring Mothers-to-Be was established to investigate the underlying causes and pathophysiological pathways associated with adverse pregnancy outcomes in nulliparous gravidas. OBJECTIVE: This study aimed to study placental physiology and identify novel biomarkers concerning adverse pregnancy outcomes, including preterm birth (medically indicated and spontaneous), preeclampsia, small-for-gestational-age neonates, and stillbirth. We measured levels of placental proteins in the maternal circulation in the first 2 trimesters of pregnancy. STUDY DESIGN: Maternal serum samples were collected at 2 study visits (6-13 weeks and 16-21 weeks), and levels of 9 analytes were measured. The analytes we measured were vascular endothelial growth factor, placental growth factor, endoglin, soluble fms-like tyrosine kinase-1, A disintegrin and metalloproteinase domain-containing protein 12, pregnancy-associated plasma protein A, free beta-human chorionic gonadotropin, inhibin A, and alpha-fetoprotein. The primary outcome was preterm birth between 20 0/7 and 36 6/7 weeks of gestation. The secondary outcomes were spontaneous preterm births, medically indicated preterm births, preeclampsia, small-for-gestational-age neonates, and stillbirth. RESULTS: A total of 10,038 eligible gravidas were enrolled in the Nulliparous Pregnancy Outcomes Study: Monitoring Mothers-to-Be cohort, from which a nested case-control study was performed comparing 800 cases with preterm birth (466 spontaneous preterm births, 330 medically indicated preterm births, and 4 unclassified preterm births), 568 with preeclampsia, 406 with small-for-gestational-age birth, and 49 with stillbirth with 911 controls who delivered at term without complications. Although levels of each analyte generally differed between cases and controls at 1 or 2 visits, the odds ratios revealed a

Histone deacetylase 6 (HDAC6) plays a crucial role in the acetylation of non-histone proteins and is notably implicated in angiogenesis, though its underlying mechanisms were previously not fully understood. This study conducted transcriptomic and proteomic analyses on vascular endothelial cells with HDAC6 knockdown, identifying endoglin (ENG) as a key downstream protein regulated by HDAC6. This protein is vital for maintaining vascular integrity and plays a complex role in angiogenesis, particularly in its interaction with bone morphogenetic protein 9 (BMP9). In experiments using human umbilical vein endothelial cells (HUVECs), the pro-angiogenic effects of BMP9 were observed, which diminished following the knockdown of HDAC6 and ENG. Western blot analysis revealed that BMP9 treatment increased SMAD1/5/9 phosphorylation, a process hindered by HDAC6 knockdown, correlating with reduced ENG expression. Mechanistically, our study indicates that HDAC6 modulates ENG transcription by influencing promoter activity, leading to increased acetylation of transcription factor SP1 and consequently altering its transcriptional activity. Additionally, the study delves into the structural role of HDAC6, particularly its CD2 domain, in regulating SP1 acetylation and subsequently ENG expression. In conclusion, the present study underscores the critical function of HDAC6 in modulating SP1 acetylation and ENG expression, thereby significantly affecting BMP9-mediated angiogenesis. This finding highlights the potential of HDAC6 as a therapeutic target in angiogenesis-related processes. [ABSTRACT FROM AUTHOR]

Hereditary hemorrhagic telangiectasia (HHT), also called Rendu–Osler syndrome, is a group of rare genetic diseases characterized by autosomal dominance, multisystemic vascular dysplasia, and age-related penetrance. This includes arteriovenous malformations (AVMs) in the skin, brain, lung, liver, and mucous membranes. The correlations between the phenotype and genotype for HHT are not clear. An HHT Chinese pedigree was recruited. Whole exome sequencing (WES) analysis, Sanger verification, and co-segregation were conducted. Western blotting was performed for monitoring ENG/VEGFα signaling. As a result, a nonsense, heterozygous variant for ENG/CD105: c.G1169A:p. Trp390Ter of the proband with hereditary hemorrhagic telangiectasia type 1 (HHT1) was identified, which co-segregated with the disease in the M666 pedigree. Western blotting found that, compared with the normal levels associated with non-carrier family members, the ENG protein levels in the proband showed approximately a one-half decrease (47.4% decrease), while levels of the VEGFα protein, in the proband, showed approximately a one-quarter decrease (25.6% decrease), implying that ENG haploinsufficiency, displayed in the carrier of this variant, may affect VEGFα expression downregulation. Pearson and Spearman correlation analyses further supported TGFβ/ENG/VEGFα signaling, implying ENG regulation in the blood vessels. Thus, next-generation sequencing including WES should provide an accurate strategy for gene diagnosis, therapy, genetic counseling, and clinical management for rare genetic diseases including that in HHT1 patients. [ABSTRACT FROM AUTHOR]

Background: Angiogenesis is the process of using existent vascular bed to form new blood vessels. Neoangiogenesis is one of the hallmarks of cancer for tumour progression and metastasis. Microvessel density (MVD) is a powerful prognostic tool in oncology for assessing the tumour vasculature. Endoglin (CD 105) is a specific marker for activated endothelium. The present research intends to investigate the prognostic and metastatic potential of Endoglin (CD 105) and immunohistochemical expression in canine squamous cell carcinoma (SCC). Methods: Totally, 15 SCC were collected from the Veterinary Clinical Complex, Tirunelveli during a one year period. The tumours were histopathologically identified as different grades. Immunohistochemically analysed with Endoglin (CD 105) and Peritumoural microvessel density and intratumourl microvessel density (PTMVD and ITMVD) were calculated using the "hot spot method". Result: All the cases were positive for CD 105 and were strongly stained with both peritumoural and intratumoural blood vessels. Among the different types of SCC, Grade III had the highest values of microvessel density. The study concluded that different grades have different MVD values within SCC cases, thereby reflecting the behaviour of neoplasms directly. Endoglin (CD 105) is one of the best endothelial markers to determine MVD because it aids to assess the prognostic and metastatic potential of tumours and it plays a significant role in the estimation of overall survival period after surgery. [ABSTRACT FROM AUTHOR]

In vitro studies have shown that Wharton's jelly mesenchymal stem cells (WJ-MSCs) can cross umbilical and uterine endothelial barriers and up-regulate endothelial junctional integrity from sub-endothelial niches. This pericytic behaviour may be lost in pregnancies complicated by gestational diabetes (GDM), where increased vascular permeability and junctional disruption are reported. The aim of the present study was to investigate whether WJ-MSCs isolated from GDM pregnancies displayed any changes in morphology, proliferation, VEGF-A secretion, and their ability to influence paracellular junctional composition and permeability. WJ-MSCs were isolated from human umbilical cords from normal pregnancies (nWJ-MSCs, n=13) and those complicated by GDM (gWJ-MSCs), either diet-controlled (d-GDM, n=13) or metformin-treated (m-GDM, n=9).We recorded that 4-fold more WJ-MSCs migrated from m-GDM, and 2.5-fold from d-GDM cord samples compared with the normal pregnancy. gWJ-MSCs showed a less predominance of spindle-shaped morphology and secreted 3.8-fold more VEGF-A compared with nWJ-MSCs. The number of cells expressing CD105 (Endoglin) was higher in gWJ-MSCs compared with nWJ-MSCs (17%) at P-2. The tracer leakage after 24 h across the HUVEC + gWJ-MSCs bilayer was 22.13% and 11.2% higher in the m-GDM and d-GDM, respectively, HUVEC + nWJ-MSCs. Transfection studies with siRNAs that target Endoglin were performed in n-WJ-MSCs; transfected cells were co-cultured with HUVEC followed by permeability studies and VE-cadherin analyses. Loss of Endoglin also led to increased VEGF-A secretion, increased permeability and affected endothelial stabilization. These results reinforce the pericytic role of nWJ-MSCs to promote vascular repair and the deficient ability of gWJ-MSCs to maintain endothelial barrier integrity. [ABSTRACT FROM AUTHOR]

Background: The increase in the collagen I (COL I)/COL III ratio enhances vessel wall stiffness and renders vessels less resistant to blood flow and pressure changes. Activated microglia enhance inflammation-induced fibrosis. Hypotheses: The COL I/COL III ratio in human and mouse brain arteriovenous malformations (bAVMs) is associated with bAVM hemorrhage, and the depletion of microglia decreases the COL I/COL III ratio and hemorrhage. Method: COL I, COL III, and hemorrhages were analyzed in 12 human bAVMs and 6 control brains, and mouse bAVMs induced in three mouse lines with activin receptor-like kinase 1 (n = 7) or endoglin (n = 7) deleted in the endothelial cells or brain focally (n = 5). The controls for the mouse study were no-gene-deleted litter mates. Mouse bAVMs were used to test the relationships between the Col I/Col III ratio and hemorrhage and whether the transient depletion of microglia reduces the Col I/Col III ratio and hemorrhage. Results: The COL I/COL III ratio was higher in the human and mouse bAVMs than in controls. The microhemorrhage in mouse bAVMs was positively correlated with the Col I/Col III ratio. Transient depletion of microglia reduced the Col I/Col III ratio and microhemorrhage. Conclusions: The COL I/COL III ratio in the bAVMs was associated with bAVM hemorrhage. The depletion of microglia reduced the bAVM Col I/Col III ratio and hemorrhage. [ABSTRACT FROM AUTHOR]

Purpose: Hereditary hemorrhagic telangiectasia (HHT) is a dominantly inherited disorder that involves epistaxis, mucocutaneous telangiectases, and visceral arteriovenous malformations (AVMs). This study aims to investigate the genetic causes in a Chinese family with HHT. Methods: HHT was confirmed according to Curaçao's diagnostic criteria. Three patients diagnosed with HHT and healthy members were recruited. Whole-exome sequencing (WES) and sanger sequencing were performed to define the patient's genetically pathogenic factor. Results: The proband presented with recurrent epistaxis, hepatopulmonary arteriovenous malformation, and adenocarcinoma. A novel frameshift mutation (c.1376_1377delAC, p.H459Lfs*41) of the ENG gene was revealed in affected individuals by WES. There was no report of this variant and predicted to be highly damaging by causing truncation of the ENG protein. Conclusion: We report a novel variant in the ENG gene in Chinese that extends the mutational and phenotypic spectra of the ENG gene, and also demonstrates the feasibility of WES in the application of genetic diagnosis of HHT. [ABSTRACT FROM AUTHOR]

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant disorder affecting 1 in 5000–8000 individuals. Hereditary hemorrhagic telangiectasia type 1 (HHT1) is the most common HHT and manifests as diverse vascular malformations ranging from mild symptoms such as epistaxis and mucosal and cutaneous telangiectases to severe arteriovenous malformations (AVMs) in the lungs, brain or liver. HHT1 is caused by heterozygous mutations in the ENG gene, which encodes endoglin, the TGFβ homodimeric co‐receptor. It was previously shown that some endoglin HHT1‐causing variants failed to traffic to the plasma membrane due to their retention in the endoplasmic reticulum (ER) and consequent degradation by ER‐associated degradation (ERAD). Endoglin is a homodimer formed in the ER, and we therefore hypothesized that mixed heterodimers might form between ER‐retained variants and WT protein, thus hampering its maturation and trafficking to the plasma membrane causing dominant negative effects. Indeed, HA‐tagged ER‐retained mutants formed heterodimers with Myc‐tagged WT endoglin. Moreover, variants L32R, V105D, P165L, I271N and C363Y adversely affected the trafficking of WT endoglin by reducing its maturation and plasma membrane localization. These results strongly suggest dominant negative effects exerted by these ER‐retained variants aggravating endoglin loss of function in patients expressing them in the heterozygous state with the WT allele. Moreover, this study may help explain some of the variability observed among HHT1 patients due to the additional loss of function exerted by the dominant negative effects in addition to that due to haploinsufficiency. These findings might also have implications for some of the many conditions impacted by ERAD. [ABSTRACT FROM AUTHOR]

We have previously demonstrated that deletion of activin receptor-like kinase 1 (Alk1) or endoglin in a fraction of endothelial cells (ECs) induces brain arteriovenous malformations (bAVMs) in adult mice upon angiogenic stimulation. Here, we addressed three related questions: (1) could Alk1- mutant bone marrow (BM)-derived ECs (BMDECs) cause bAVMs? (2) is Alk1- ECs clonally expended during bAVM development? and (3) is the number of mutant ECs correlates to bAVM severity? For the first question, we transplanted BM from PdgfbiCreER;Alk12f/2f mice (EC-specific tamoxifen-inducible Cre with Alk1-floxed alleles) into wild-type mice, and then induced bAVMs by intra-brain injection of an adeno-associated viral vector expressing vascular endothelial growth factor and intra-peritoneal injection of tamoxifen. For the second question, clonal expansion was analyzed using PdgfbiCreER;Alk12f/2f;confetti+/- mice. For the third question, we titrated tamoxifen to limit Alk1 deletion and compared the severity of bAVM in mice treated with low and high tamoxifen doses. We found that wild-type mice with PdgfbiCreER;Alk12f/2f BM developed bAVMs upon VEGF stimulation and Alk1 gene deletion in BMDECs. We also observed clusters of ECs expressing the same confetti color within bAVMs and significant proliferation of Alk1- ECs at early stage of bAVM development, suggesting that Alk1- ECs clonally expanded by local proliferation. Tamoxifen dose titration revealed a direct correlation between the number of Alk1- ECs and the burden of dysplastic vessels in bAVMs. These results provide novel insights for the understanding of the mechanism by which a small fraction of Alk1 or endoglin mutant ECs contribute to development of bAVMs.

Endoglin (ENG) mutation causes type 1 hereditary hemorrhagic telangiectasia (HHT1). HHT1 patients have arteriovenous malformations (AVMs) in multiple organs, including the brain. In mice, Eng deletion induced by R26RCreER or SM22αCre leads to AVM development in the brain and other organs. We hypothesized that an increase in Eng- negative ECs will enhance AVM severity. To increase EC Eng deletion, we used a codon-improved cre (icre), which is more potent in recombination of the floxed alleles than the wild-type (WT) cre. R26RCreER;Engf/f mice that have a Rosa promoter driving and tamoxifen (TM)-inducible WT cre expression globally, and PdgfbiCreER;Engf/f mice that have a Pdgfb promoter driving and TM-inducible icre expression in ECs were treated with three intra-peritoneal injections of TM (2.5 mg/25 g of body weight) to delete Eng globally or in the ECs. AAV-VEGF was stereotactically injected into the brain to induce brain focal angiogenesis and brain AVM. We found that icre caused more Eng deletion in the brain, indicated by a lower level of Eng proteins (p < 0.001) and fewer Eng-positive ECs (p = 0.01) than mice with WT cre. Mice with icre-mediated Eng deletion have more abnormal vessels (p = 0.02), CD68+ macrophages (p = 0.002), and hemorrhage (p = 0.04) and less vascular pericyte and smooth muscle coverage than mice with WT cre. In addition, arteriovenous shunts were detected in the intestines of icre mice, a phenotype that has not been detected in WT cre mice before. RNA-seq analysis showed that 8 out of the 10 top upregulated pathways identified by gene ontology (GO) analysis are related to inflammation. Therefore, the increase in Eng deletion in ECs exacerbates AVM severity, which is associated with enhanced inflammation. Strategies that can reduce Eng-negative ECs could be used to develop new therapies to reduce AVM severity for HHT1 patients.

BackgroundExtracellular vehicles (EVs) released by malignant tumor cells can mediate the immune response and promote metastasis through intercellular communication. EV analysis is an emerging cancer surveillance tool with advantages over traditional liquid biopsy methods. The aim of this pilot study is to identify actionable EV signatures in metastatic breast cancer.Materials and methodsUnder an IRB-approved protocol for the analysis of patient plasma, samples were collected from women with newly diagnosed or progressive metastatic breast cancer and from women without cancer. Enriched EVs were analyzed via a bead-based multiplex assay designed to detect 37 distinct tumor-relevant epitopes. The mean fluorescent intensity of EV epitopes meeting a minimum threshold of detectability was compared between groups via independent samples t-test. Subgroup analysis was conducted for metastatic breast cancer patients who were positive for estrogen and/or progesterone receptors and negative for HER2. Other variables potentially affecting CD105 levels were also analyzed.ResultsCD105 was found to have a significantly higher mean fluorescent intensity in participants with metastatic breast cancer compared to control participants (P = 0.04). ER/PR+ subgroup analysis revealed a similar pattern compared to control participants (P = 0.01). Other analyzed variables were not found to have a significant correlation with CD105 levels.ConclusionsCD105 EV levels were significantly higher in samples from participants with breast cancer compared to controls. Given that CD105 is known to mediate angiogenesis and promote metastasis, EV-associated CD105 in plasma represents a potential biomarker for diagnosis, surveillance and therapeutic targeting in patients with metastatic breast cancer.

BackgroundRetrospective questionnaire and healthcare administrative data suggest reduced life expectancy in untreated hereditary hemorrhagic telangiectasia (HHT). Prospective data suggests similar mortality, to the general population, in Denmark's centre-treated HHT patients. However, clinical phenotypes vary widely in HHT, likely affecting mortality. We aimed to measure predictors of mortality among centre-treated HHT patients. HHT patients were recruited at 14 HHT centres of the Brain Vascular Malformation Consortium (BVMC) since 2010 and followed annually. Vital status, organ vascular malformations (VMs) and clinical symptoms data were collected at baseline and during follow-up (N = 1286). We tested whether organ VMs, HHT symptoms and HHT genes were associated with increased mortality using Cox regression analysis, adjusting for patient age, sex, and smoking status.Results59 deaths occurred over average follow-up time of 3.4 years (max 8.6 years). A history of anemia was associated with increased mortality (HR = 2.93, 95% CI 1.37-6.26, p = 0.006), as were gastro-intestinal (GI) bleeding (HR = 2.63, 95% CI 1.46-4.74, p = 0.001), and symptomatic liver VMs (HR = 2.10, 95% CI 1.15-3.84, p = 0.015). Brain VMs and pulmonary arteriovenous malformations (AVMs) were not associated with mortality (p > 0.05). Patients with SMAD4 mutation had significantly higher mortality (HR = 18.36, 95% CI 5.60-60.20, p

BackgroundApproximately 10% of hereditary hemorrhagic telangiectasia (HHT) patients harbour brain vascular malformations (VMs). Intracranial hemorrhage (ICH) from brain VMs can lead to death or morbidity, while treatment options for brain VMs also have associated morbidity. The modified Rankin Scale (mRS) may provide an approach to identifying HHT-brain VM patients with poor outcomes, and their predictors. We aimed to measure the relationship between mRS score and brain VM, brain VM number, as well as other aspects of HHT, at enrollment and during prospective follow-up.Methods1637 HHT patients (342 with brain VMs) were recruited from 14 HHT centres of the Brain Vascular Malformation Consortium since 2010 and followed prospectively (mean = 3.4 years). We tested whether the presence of brain VM, other HHT organ involvement, and HHT mutation genotype were associated with worse mRS scores at baseline and during follow-up, using linear mixed models, adjusting for age, sex, and year of visit.ResultsPresence of brain VMs was not associated with worse mRS score at baseline and there was no significant worsening of mRS with prospective follow-up in these patients; 92% had baseline mRS of 0-2. HHT-related gastrointestinal (GI) bleeding was associated with worse mRS scores at baseline (0.37, 95% CI 0.26-0.47, p

One of the most important characteristics of radiolabeled antibodies and their derivatives is the size of the immunoreactive fraction. Measuring this parameter requires a high density of target molecules, which is rarely achievable with tumor cells. The solution to the problem of radioimmunoconjugate testing was the creation of recombinant cells carrying human endoglin (CD105). The recipients of the endoglin gene (ENG) were rat C6 glioma cells, which are characterized by ease of cultivation and high transfection efficiency. The obtained C6-ENG cells carried 1.3 × 106 CD105 molecules on the membrane and were used to determine the immunoreactive fraction of 68Ga and 89Zr radiolabeled anti-CD105 monoclonal antibodies and their Fab-fragments. The creation of stable recombinant cell lines for in vitro testing the specific activity of radiolabeled antibodies and their derivatives seems promising for the development of new radiopharmaceuticals. [ABSTRACT FROM AUTHOR]

Brain arteriovenous malformations (BAVMs) are a critical concern in hereditary hemorrhagic telangiectasia (HHT) patients, carrying the risk of life-threatening intracranial hemorrhage. While traditionally seen as congenital, the debate continues due to documented de novo cases. Our primary goal was to identify the precise postnatal window in which deletion of the HHT gene Endoglin (Eng) triggers BAVM development. We employed SclCreER(+);Eng2f/2f mice, enabling timed Eng gene deletion in endothelial cells via tamoxifen. Tamoxifen was given during four postnatal periods: P1–3, P8–10, P15–17, and P22–24. BAVM development was assessed at 2–3 months using latex dye perfusion. We examined the angiogenic activity by assessing vascular endothelial growth factor receptor 2 (VEGFR2) expression via Western blotting and Flk1-LacZ reporter mice. Longitudinal magnetic resonance angiography (MRA) was conducted up to 9 months. BAVMs emerged in 88% (P1–3), 86% (P8–10), and 55% (P15–17) of cases, with varying localization. Notably, the P22–24 group did not develop BAVMs but exhibited skin AVMs. VEGFR2 expression peaked in the initial 2 postnatal weeks, coinciding with BAVM onset. These findings support the "second hit" theory, highlighting the role of early postnatal angiogenesis in initiating BAVM development in HHT type I mice. [ABSTRACT FROM AUTHOR]

TGF-β is recognized as playing a protective role against atherosclerosis. Endoglin is a receptor for TGF-β, and its expression is upregulated in atherosclerotic plaques. Endoglin is secreted from the cell membrane into the circulation as a soluble form (sEng). We previously reported that plasma sEng levels were low in patients with coronary artery disease (CAD). However, the prognostic value of sEng levels has not been clarified. We investigated the association between plasma sEng levels and cardiovascular events in 403 patients who had an elective coronary angiography and were then followed up. Cardiovascular events were defined as cardiovascular death, myocardial infarction, unstable angina, heart failure, stroke, or coronary revascularization. Of the 403 patients, 209 (52%) had CAD. Plasma sEng levels were lower in patients with CAD than in those without CAD (median 4.26 vs. 4.41 ng/mL, p < 0.025). During a mean follow-up period of 7.5 ± 4.5 years, cardiovascular events occurred in 79 patients. Compared with 324 patients without events, 79 with events had lower sEng levels (3.95 vs. 4.39 ng/mL) and more often had an sEng level < 3.9 ng/mL (47% vs. 28%) (p < 0.02). A Kaplan–Meier analysis showed lower event-free survival in patients with sEng < 3.9 ng/mL than in those with ≥3.9 ng/mL (p < 0.02). In a multivariate Cox proportional hazards analysis, the sEng level (<3.9 ng/mL) was an independent predictor of cardiovascular events (hazard ratio: 1.59; 95%CI: 1.01–2.49). Furthermore, only among the 209 patients with CAD, the sEng level was also a predictor of further cardiovascular events (hazard ratio: 2.07; 95%CI: 1.24–3.45). Thus, low plasma sEng levels were found to be associated with an increased risk of cardiovascular events in patients with CAD and patients undergoing coronary angiography. [ABSTRACT FROM AUTHOR]

Background: The aim of the present study is to determine the possibility of isolation and characterization of the human periodontal ligament stem cells (hPDLSCs) using limited harvested periodontal ligament (PDL) tissue of only one patient's wisdom teeth (2--4 teeth) under the more compatible terms of use in clinical application without using the fetal bovine serum (FBS). Materials and Methods: In this pilot study, hPDLSCs were isolated from the impacted third molar, and tissue was scraped from the roots of the impacted third molar of 10 volunteers to enzymatically digest using collagenase. The cells were sub-cultured. The samples of the first seven patients and half of the eighth patient's sample were cultured in alpha modified of Eagle's medium (α-MEM) (FBS) medium and the other part of the eighth patient's sample was cultured with prior medium supplemented with +FBS 15% as a control of the cultivation protocol. While for the past two patients (9th and 10th the α-MEM medium was supplemented with L-Glutamine, anti/anti 2X, and 20% knock-out serum replacement (KSR). Two more nutritious supplements (N2 and B27) were added to the medium of the tenth sample. Flow-cytometric analysis for the mesenchymal stem cell surface markers CD105, CD45, CD90, and CD73 was performed. Subsequent polymerase chain reaction was undertaken on three samples cultured with two growth media. Results: Cultivation failed in some of the samples because of the lack of cell adhesion to the culturing dish bottom (floating cells), but it was successful for the 9th and 10th patients, which were cultured in the α-MEM serum supplemented with KSR 20%. Flow cytometry analysis was positive for CD105, CD90, and CD73 and negative for CD45. The PDL stem cells (PDLSCs) expressed CD105, CD45, and CD90 but were poor for CD73. Conclusion: According to the limited number of sample tests in this study, isolation and characterization of PDLSCs from collected PDL tissue of one patient's wisdom teeth (2--4) may be possible by the proper setup in synthetic FBS-free serum. [ABSTRACT FROM AUTHOR]

This study aimed to evaluate the clinicopathologic significance of the combined immunohistochemical expression of the epithelial-mesenchymal transition marker E-cadherin and the angiogenesis marker CD105 in laryngeal squamous cell carcinoma and assess correlation of their expression. Eighty-five patients who underwent complete resection as primary treatment were selected for this study. E-cadherin and CD105 expression levels were determined by immunohistochemistry. The receiver operating curve approach was applied to determine the cut-off value and separate patients with high and low expression of markers. The high-risk group ("CD105 high" and "E-cadherin low" expression) showed statistically significant correlations with age less than 66 years (p = 0.039), advanced T-status (T3-4) (p = 0.046), aggressive TNM stage (stage III--IV) (p = 0.003) and loco-regional recurrence of disease (p = 0.004). In the Kaplan-Meier analyses, the high-risk group had significantly worse prognoses than other risk groups (log-rank test χ² = 9.415, p = 0.024). Spearman's correlation coefficient analysis showed a nonsignificant negative correlation between the expression of E-cadherin and CD105 (rho = --0.073, p = 0.505). Simultaneous consideration of E-cadherin and CD105 is a simple panel of markers to determine aggressive tumour phenotype with a higher risk of disease recurrence in patients with laryngeal cancer. [ABSTRACT FROM AUTHOR]

Lessons learnedThe combination of carotuximab with axitinib did not provide a benefit over axitinib monotherapy in patients with metastatic clear cell renal cell carcinoma who had previously progressed on one or more vascular endothelial growth factor (VEGF)-targeted therapies. Exploratory evaluation of pretreatment circulating biomarkers suggested the combination might benefit patients who have low baseline VEGF levels.BackgroundEndoglin is an angiogenic receptor expressed on proliferating tumor vessels and renal cell carcinoma (RCC) stem cells that is implicated as a mechanism of resistance to vascular endothelial growth factor receptor (VEGFR) inhibitors. This study evaluated an antiendoglin monoclonal antibody (carotuximab, TRC105) combined with axitinib in patients with advanced or metastatic clear cell renal cell carcinoma (mccRCC) who had progressed following one or more prior VEGF inhibitors.MethodsTRAXAR was a multicenter, international randomized 1:1 (stratified by ECOG, 0 vs. 1), phase II study of carotuximab combined with axitinib versus axitinib alone in mccRCC patients who had progressed following one or more prior VEGF inhibitors. The primary endpoint was progression-free survival (PFS) assessed by independent central review (ICR) per RECIST 1.1 RESULTS: A total of 150 patients were randomized. The combination therapy resulted in shorter median PFS by RECIST 1.1 than axitinib monotherapy (6.7 vs. 11.4 months). The combination was tolerated similarly to axitinib monotherapy, and there were no treatment related deaths. Exploratory evaluation of pretreatment circulating biomarkers suggested the combination might benefit patients who have low baseline VEGF levels.ConclusionThe combination of carotuximab with axitinib did not demonstrate additional efficacy over single agent axitinib in patients with mccRCC who progressed following one or more prior VEGF inhibitor treatment.

BACKGROUND: Primary hyperparathyroidism (PHPT), one of the most common endocrine pathologies, is associated with a higher incidence of cardiovascular diseases, in particular, those caused by endothelial dysfunction. Evaluation of endothelial dysfunction in patients with PHPT will predict the development of cardiovascular pathology and determine the optimal tactics for PHPT management.AIM: To evaluate the concentration of soluble endoglin and photoplethysmographic parameters as potential markers of endothelial dysfunction in patients with PHPT.MATERIALS AND METHODS: A single-center interventional single-stage study was carried out. 2 groups were formed. The first group included 50 patients with verified PHPT who did not have cardiovascular or other concomitant somatic pathologies in anamnesis. The comparison group included 21 healthy volunteers comparable in sex and age. All participants underwent a biochemical blood test (total calcium, ionized, albumin, lipidogram, urea, uricacid, glucose, creatinine, alkaline phosphatase), parathyroid hormone, 25 (OH) D and endoglin concentrations were evaluated. In addition, echocardiography, ultrasound of the brachiocephalic arteries and arteries of the lower extremities, as well as photoplethysmography were performed.RESULTS: The groups differed in mineral parameters associated with PHPT; no differences were found in parameters of lipid, uric acid and carbohydrate metabolism. Serum levels of endoglin were lower in PHPT patients (p=0.002). We found a negative correlation between the concentration of albumin-corrected calcium and PTH with endoglin (r1=-0.370, p1=0.003 and r2=-0.475, p2

Chimeric antigen receptor-based T-cell immunotherapy is a promising strategy for treatment of hematological malignant tumors; however, its efficacy towards solid cancer remains challenging. We therefore focused on developing nanobody-based CAR-T cells that treat the solid tumor. CD105 expression is upregulated on neoangiogenic endothelial and cancer cells. CD105 has been developed as a drug target. Here we show the generation of a CD105-specific nanobody, an anti-human CD105 CAR-T cells, by inserting the sequences for anti-CD105 nanobody-linked standard cassette genes into AAVS1 site using CRISPR/Cas9 technology. Co-culture with CD105+ target cells led to the activation of anti-CD105 CAR-T cells that displayed the typically activated cytotoxic T-cell characters, ability to proliferate, the production of pro-inflammatory cytokines, and the specific killing efficacy against CD105+ target cells in vitro. The in vivo treatment with anti-CD105 CAR-T cells significantly inhibited the growth of implanted CD105+ tumors, reduced tumor weight, and prolonged the survival time of tumor-bearing NOD/SCID mice. Nanobody-based CAR-T cells can therefore function as an antitumor agent in human tumor xenograft models. Our findings determined that the strategy of nanobody-based CAR-T cells engineered by CRISPR/Cas9 system has a certain potential to treat solid tumor through targeting CD105 antigen.

Purpose: To determine whether 14 inflammation-, angiogenesis-, and adhesion-related proteins in cord blood (CB), alone or in combination with conventional perinatal factors, could predict retinopathy of prematurity (ROP) in preterm infants. Methods: Data from 111 preterm infants (born at ≤ 32.0 weeks) were retrospectively reviewed. The levels of endoglin, E-selectin, HSP70, IGFBP-3/4, LBP, lipocaline-2, M-CSFR, MIP-1α, pentraxin 3, P-selectin, TGFBI, TGF-β1, and TNFR2 were assessed in stored CB samples collected at birth using ELISA kits. The primary endpoints included severe ROP (≥ stage 3) and type 1 ROP requiring treatment. Results: ROP was diagnosed in 29 infants (26.1%), among whom 14 (12.6%) had severe ROP and seven (6.3%) had type 1 ROP. Multivariate logistic regression showed that decreased CB TGFBI levels were significantly associated with severe ROP and type 1 ROP after adjusting for gestational age at birth. Stepwise regression analysis allowed to design prediction models with good accuracy, which comprised low CB TGFBI levels and low birth weight (BW) as predictors for severe ROP (area under the curve [AUC] = 0.888), and low CB endoglin levels and low BW as predictors for type 1 ROP (AUC = 0.950). None of the other CB proteins evaluated were found to be associated with severe ROP or type 1 ROP. Conclusions: Low CB TGFBI levels are associated with severe ROP and type 1 ROP, independently of gestational age. Moreover, combined predictive models based on CB TGFBI and endoglin levels, along with BW data, may act as good indicators at birth for the neonatal risk of ROP progression. [ABSTRACT FROM AUTHOR]

An unbalance of angiogenesis, including soluble endoglin, exists that causes preeclampsia. Preeclampsia patients had considerably higher serum soluble endoglin levels compared to women who were not experiencing any complications during their pregnancies. Comparing the serum concentrations of soluble endoglin between pregnant women experiencing preeclampsia and those who had normal blood pressure. Forty pregnant women who were at least 20 weeks along were split evenly into two groups: those with preeclampsia and those without. Venipuncture was used to collect 5ml of blood into a test tube without the use of anticoagulant. Kits were used to perform a one-step sandwich enzyme immunoassay on plasma endoglin with monoclonal antibodies specific to human endoglin. The blood soluble endoglin level was significantly greater in the preeclampsia group (18.52 9.54 vs. 2.2 1.4, p= 0.000), and there was positive association between an elevated serum endoglin level and the probability of pre-eclampsia (r=0.523, P=0.016). The optimal sensitivity and specificity was found at the former was greater whereas the latter was lower than the threshold value of 6.26500 ng/ml.The diagnostic accuracy of serum soluble endoglin for preeclampsia is exceptional. [ABSTRACT FROM AUTHOR]

Introduction. In this study we analyzed CD105 (endoglin) and E-cadherin expression in laryngeal squamous cell carcinoma (LSCC) to evaluate their clinicopathologic significance. Material and methods. Expression of CD105 and E-cadherin was examined immunohistochemically using paraffin- -embedded archival tissues of 72 (35 glottic and 37 supraglottic) previously untreated LSCC male patients. The mean value of the positively-stained microvessels for CD105 counted in four hot spots for each case was used as the final intratumoralmicrovessel density (MVD). A staining score of E-cadherin was calculated based on the percentage of cells stained (0--100%). Results. MVD was significantly higher in patients with advanced TNM stage (P = 0.004) and younger than 65 (P = 0.008). Nodal metastases were more frequent in the cases with low E-cadherin expression (P = 0.000). Tumor recurrence was associated with advanced TNM stage (P = 0.035) and high MVD (P = 0.002). A high MVD was an independent predictor of malignancy recurrence (P = 0.021). The log-rank test showed a significant difference in the disease-free interval in patients stratified according to the MVD value (P = 0.016). Spearman's rank correlation test did not show a significant correlation between E-cadherin and CD105 expression. Conclusions. CD105-assessed MVD and expression of E-cadherin are promising prognostic factors for the outcome of patients with LSCC. Increased expression of CD105 could help predict patients with an increased risk of developing loco-regional recurrence after surgical treatment. Decreased E-cadherin expression is a potential predictor of lymph node metastases. (Folia Histochemica et Cytobiologica 2023, Vol. 61, No. 3, 183--192) [ABSTRACT FROM AUTHOR]

Islet transplantation into subcutaneous polymer scaffolds has shown to successfully induce normoglycemia in type 1 diabetes models. Vascularization of these scaffolds is imperative for optimal control of glucose levels. We studied the effect of the vascular stimulator hydrogen sulfide (H2S) on the degree of vascularization of a scaffold and the role of the immune system in this process. Scaffolds were subcutaneously implanted in immunocompetent C57BL/6 and immunocompromised nude mice. Mice received twice-daily intraperitoneal injections of the fast-releasing H2S donor sodium hydrosulfide (NaHS, 25 or 50 μmol/kg) or saline for 28 days. After 63 days the vascular network was analyzed by histology and gene expression. Here we showed that the vascularization of a subcutaneous scaffold in nude mice was significantly impaired by H2S treatment. Both the CD31 gene and protein expression were reduced in these scaffolds compared to the saline-treated controls. In C57BL/6 mice, the opposite was found, the vascularization of the scaffold was significantly increased by H2S. The mRNA expression of the angiogenesis marker CD105 was significantly increased compared to the controls as well as the number of CD31 positive blood vessels. In conclusion, the immune system plays an important role in the H2S mediated effect on vascularization of subcutaneous scaffolds.

Background Diabetic retinopathy (DR) signifies a frequent serious diabetic complication influencing retinal structure and function. Dysregulation of lncRNAs drives a wide array of human diseases especially diabetes; thus, we aimed to study lncRNA HIF1A-AS2 role and its interplay with hypoxia, oxidative stress (OS), and angiogenesis in DR.Materials and methods 60 DM patients in addition to 15 healthy subjects. were enrolled. LncRNA HIF1A-AS2 mRNA relative gene expression was assessed. Hypoxia inducible factor 1-alpha (HIF-1α), vascular endothelial growth factor (VEGF), mitogen activated protein kinase (MAPK), and endoglin levels were assessed. Detection of DNA damage using comet assay, and Redox status parameters were also detected.Results LncRNA HIF1A-AS2 expression was significantly increased in diabetic patients with the highest levels in proliferative DR patients. Moreover, HIFα, VEGF, MAPK, and Endogolin levels were significantly higher in the diabetic patients compared to control group with the highest levels in in proliferative DR patients. Significant DNA damage in comet assay was observed to be the highest in this group.Conclusion We observed for the first time the imminent role of long noncoding RNA HIF1A-AS2 in DR throughout its stages and its interplay with hypoxia, OS, and angiogenesis via MAPK/VEGF-dependent pathway.

Purpose To evaluate the level of soluble endoglin (sEng) in pregnant women with pregestational diabetes mellitus (DM) and to assess its predictive value for preeclampsia development. Methods Ninety pregnant women were enrolled in the study forming five comparison groups: type 1 DM (not planned, n = 20; planned, n = 20), type 2 DM (diet, n = 15; insulin therapy, n = 20), and the control group (n = 15). The primary outcome was clinically confirmed preeclampsia. Maternal serum concentrations of sEng were measured at 11+0–13+6 and 30+0–33+6 weeks. Results sEng level was elevated in all patients with pregestational DM compared to the control group. Its plasma concentration increased with gestational age and in case of preeclampsia development. In patients with type 1 DM, serum sEng level did not depend on the presence of preeclampsia. This is evidence of severe metabolic disorder and endothelial dysfunction in these patients. The addition of sEng level to logistic models considering established risk factors (body mass index + age + HbA1c level) in the first and third trimesters significantly improved their performance for preeclampsia prediction. Conclusions Eng level may become an important marker for early prediction of preeclampsia in women with pregestational DM.

Myocytic tumors of the uterus present vast morphological heterogeneity, which makes differential diagnosis between the different entities necessary. This study aims to enrich the existing data and highlight new potential therapeutic targets regarding aspects related to the pathogenic process and the tumor microenvironment in order to improve the quality of life of women. We performed a 5-year retrospective study, including particular cases of uterine myocyte tumors. Immunohistochemical analyses of pathogenic pathways (p53, RB1, and PTEN) and tumor microclimate using markers (CD8, PD-L1, and CD105), as well as genetic testing of the PTEN gene, were performed. The data were statistically analyzed using the appropriate parameters. In cases of atypical leiomyoma, a significant association was observed between PTEN deletion and an increased number of PD-L1+ T lymphocytes. For malignant lesions and STUMP, PTEN deletion was associated with the advanced disease stage. Advanced cases were also associated with an increased mean CD8+ T cell count. An increased number of lymphocytes was associated with an increased percentage of RB1+ nuclei. The study corroborated clinical and histogenetic data, highlighting the importance of the differential diagnosis of these tumors to improve the management of patients and increase their quality of life. [ABSTRACT FROM AUTHOR]

CD105 (endoglin) is a transmembrane protein that functions as a TGF-beta coreceptor and is highly expressed on endothelial cells. Unsurprisingly, preclinical and clinical evidence strongly suggests that CD105 is an important contributor to tumor angiogenesis and tumor progression. Emerging evidence suggests that CD105 is also expressed by tumor cells themselves in certain cancers such as renal cell carcinoma (RCC). In human RCC tumor cells, CD105 expression is associated with stem cell-like properties and contributes to the malignant phenotype in vitro and in xenograft models. However, as a regulator of TGF-beta signaling, there is a striking lack of evidence for the role of tumor-expressed CD105 in the anti-tumor immune response and the tumor microenvironment. In this study, we report that tumor cell-expressed CD105 potentiates both the in vitro and in vivo tumorigenic potential of RCC in a syngeneic murine RCC tumor model. Importantly, we find that tumor cell-expressed CD105 sculpts the tumor microenvironment by enhancing the recruitment of immunosuppressive cell types and inhibiting the polyfunctionality of tumor-infiltrating CD4+ and CD8+ T cells. Finally, while CD105 expression by endothelial cells is a well-established contributor to tumor angiogenesis, we also find that tumor cell-expressed CD105 significantly contributes to tumor angiogenesis in RCC. [ABSTRACT FROM AUTHOR]

Angiogenesis is an essential part of normal skin healing, re‐establishing blood flow in developing granulation tissue. Non‐healing skin wounds are associated with impaired angiogenesis and although the role of re‐establishing macroscopic blood flow to limbs to prevent wound chronicity is well investigated, less is known about vascular alterations at the microcirculatory level. We hypothesised that significant phenotypic changes would be evident in blood vessels surrounding chronic skin wounds. Wound edge tissue, proximal to wound (2 cm from wound edge) and non‐involved skin (>10 cm from wound edge) was harvested under informed consent from 20 patients undergoing elective amputation due to critical limb ischemia. To assess blood vessel structure and viability, tissue was prepared for histological analysis and labelled with antibodies specific for PECAM‐1 (CD31), CD146, endoglin, ALK‐1, ALK‐5, and p16Ink4a as a marker of cellular senescence. Density of microvasculature was significantly increased in wound edge dermis, which was concomitant with increased labelling for endoglin and CD146. The number of CD31 positive vessel density was unchanged in wound edge tissue relative to non‐involved tissue. Co‐labelling of endoglin with the transforming growth factor receptor ALK‐1, and to a lesser extent ALK‐5, demonstrated activation of endothelial cells which correlated with PCNA labelling indicative of proliferation. Analysis of p16Ink4a staining showed a complete lack of immunoreactivity in the vasculature and dermis, although staining was evident in sub‐populations of keratinocytes. We conclude that the endoglin‐ALK‐1‐endothelial proliferation axis is active in the vasculature at the edge of chronic skin wounds and is not associated with p16Ink4a mediated senescence. This information could be further used to guide treatment of chronic skin wounds and optimise debridement protocols. [ABSTRACT FROM AUTHOR]

Angiogenesis is an essential part of normal skin healing, re‐establishing blood flow in developing granulation tissue. Non‐healing skin wounds are associated with impaired angiogenesis and although the role of re‐establishing macroscopic blood flow to limbs to prevent wound chronicity is well investigated, less is known about vascular alterations at the microcirculatory level. We hypothesised that significant phenotypic changes would be evident in blood vessels surrounding chronic skin wounds. Wound edge tissue, proximal to wound (2 cm from wound edge) and non‐involved skin (>10 cm from wound edge) was harvested under informed consent from 20 patients undergoing elective amputation due to critical limb ischemia. To assess blood vessel structure and viability, tissue was prepared for histological analysis and labelled with antibodies specific for PECAM‐1 (CD31), CD146, endoglin, ALK‐1, ALK‐5, and p16Ink4a as a marker of cellular senescence. Density of microvasculature was significantly increased in wound edge dermis, which was concomitant with increased labelling for endoglin and CD146. The number of CD31 positive vessel density was unchanged in wound edge tissue relative to non‐involved tissue. Co‐labelling of endoglin with the transforming growth factor receptor ALK‐1, and to a lesser extent ALK‐5, demonstrated activation of endothelial cells which correlated with PCNA labelling indicative of proliferation. Analysis of p16Ink4a staining showed a complete lack of immunoreactivity in the vasculature and dermis, although staining was evident in sub‐populations of keratinocytes. We conclude that the endoglin‐ALK‐1‐endothelial proliferation axis is active in the vasculature at the edge of chronic skin wounds and is not associated with p16Ink4a mediated senescence. This information could be further used to guide treatment of chronic skin wounds and optimise debridement protocols. [ABSTRACT FROM AUTHOR]

Background: Pyogenic granuloma (PG) is one of the most prevalent tumor-like lesions in the mouth, with two distinct histological variants: Non-Lobular Capillary Hemangioma (NLCH) and Lobular Capillary Hemangioma (LCH). This study aims to evaluate and compare the immunohistochemical expression of Ki-67 (proliferative activity) and CD 105 (angiogenic activity) in both types to find the conflict between the two. Methods: Data from clinical information and microscopic findings were collected. The selection criteria, including sufficient tissue, minor bleeding and/or inflammation, and proper fixation, were used to examine 24 cases of PG (10 LCH, 14 NLCH) that performed hematoxylin and eosin (H&E) staining and immunostaining of anti-Ki-67 antibodies and anti-CD105 antibodies. Results: Our study showed female predilection, as 66.7% of cases were female (10 cases in LCH and 6 cases in NLCH). The mean patient age was 32.71%±15.35 years. LCH type of PG showed moderate positive expression of Ki-67 in 9 cases (64.3%); only two cases (14.3%) had a negative expression, while in 6 cases (60%) of NLCH, mild positive expression was seen, and in 4 cases (40%) it was moderate. The average number of blood vessels measured in LCH and NLCH samples treated with the anti-CD105 antibody was (72.7857±39.21479) and (47.1600±30.02855) respectively, with no significant difference between the two types of PG for both markers. Conclusions: The results of the present study confirm that there are no differences between the LCH and NLCH types of PG in proliferative rate and angiogenesis activity, so both types seem to have the same biological behavior. [ABSTRACT FROM AUTHOR]

Endoglin (ENG) is a single-pass transmembrane protein highly expressed on vascular endothelial cells, although low expression levels can be detected in many other cell types. Its extracellular domain can be found in circulation known as soluble endoglin (sENG). Levels of sENG are elevated in many pathological conditions, in particular preeclampsia. We have shown that while loss of cell surface ENG decreases BMP9 signaling in endothelial cells, knocking down ENG in blood cancer cells enhances BMP9 signaling. Despite sENG binding to BMP9 with high affinity and blocking the type II receptor binding site on BMP9, sENG did not inhibit BMP9 signaling in vascular endothelial cells, but the dimeric form of sENG inhibited BMP9 signaling in blood cancer cells. Here we report that in non-endothelial cells such as human multiple myeloma cell lines and the mouse myoblast cell line C2C12, both monomeric and dimeric forms of sENG inhibit BMP9 signaling when present at high concentrations. Such inhibition can be alleviated by the overexpression of ENG and ACVRL1 (encoding ALK1) in the non-endothelial cells. Our findings suggest that the effects of sENG on BMP9 signaling is cell-type specific. This is an important consideration when developing therapies targeting the ENG and ALK1 pathway. [ABSTRACT FROM AUTHOR]