TGF-β1 Expression Research Articles

Exploring dysfunctional barrier phenotypes associated with glaucoma using a human pluripotent stem cell-based model of the neurovascular unit

AbstractGlaucoma is a neurodegenerative disease that results in the degeneration of retinal ganglion cells (RGCs) and subsequent loss of vision. While RGCs are the primary cell type affected in glaucoma, neighboring cell types selectively modulate RGCs to maintain overall homeostasis. Among these neighboring cell types, astrocytes, microvascular endothelial cells (MVECs), and pericytes coordinate with neurons to form the neurovascular unit that provides a physical barrier to limit the passage of toxic materials from the blood into neural tissue. Previous studies have demonstrated that these barrier properties may be compromised in the progression of glaucoma, yet mechanisms by which this happens have remained incompletely understood. Thus, the goals of this study were to adapt a human pluripotent stem cell (hPSC)-based model of the neurovascular unit to the study of barrier integrity relevant to glaucoma. To achieve this, hPSCs were differentiated into the cell types that contribute to this barrier, including RGCs, astrocytes, and MVECs, then assembled into an established Transwell®-insert model. The ability of these cell types to contribute to an in vitro barrier model was tested for their ability to recapitulate characteristic barrier properties. Results revealed that barrier properties of MVECs were enhanced when cultured in the presence of RGCs and astrocytes compared to MVECs cultured alone. Conversely, the versatility of this system to model aspects of barrier dysfunction relevant to glaucoma was tested using an hPSC line with a glaucoma-specific Optineurin (E50K) mutation as well as a paired isogenic control, where MVECs then exhibited reduced barrier integrity. To identify factors that could result in barrier dysfunction, results revealed an increased expression of TGFβ2 in glaucoma-associated OPTN(E50K) astrocytes, indicating a potential role for TGFβ2 in disease manifestation. To test this hypothesis, we explored the ability to modulate exogenous TGFβ2 in both isogenic control and OPTN(E50K) experimental conditions. Collectively, the results of this study indicated that the repurposing of this in vitro barrier model for glaucoma reliably mimicked some aspects of barrier dysfunction, and may serve as a platform for drug discovery, as well as a powerful in vitro model to test the consequences of barrier dysfunction upon RGCs in glaucoma.

Dietary intervention of propolis and/or turmeric boosted growth, hematology, biochemical profile, and antioxidant-immune responses and their associated gene expression in Nile tilapia (Oreochromis niloticus) challenged with Edwardsiella tarda

AbstractImmunostimulant phytogenic feed additives are given great concern for improving fish health, growth, immune responses, and resistance to diseases. This research investigated the impact of dietary propolis (PRO), turmeric (TUR), and their combination on the growth, hematology, antioxidant-immune responses, and their regulating genes in Nile tilapia (Oreochromis niloticus) during Edwardsiella tarda challenge. For 8 weeks, a total number of 320 fish (20.70 ± 0.14 g) were allocated into four groups at random, each with eight replicates (10 fish each). The first group (1st) was given the basal diet (control) without any supplements. The 2nd, 3rd, and 4th groups were supplemented with 1% PRO, 1% TUR, and the mixture (1%PRO + 1%TUR), respectively. The experimental groups were challenged intraperitoneally with E. tarda at a dose of 0.1 mL (1 × 105 CFU) at the termination of the feeding trial, and the fish survival was estimated for an additional 7 days. The results demonstrated that fish-fed diets supplemented with PRO and/or TUR showed higher body weight, condition factor, specific growth rate, feed intake, and feed efficiency utilization than the control group (P < 0.05). The hematological, protein profile, and antioxidant-immune (total antioxidant capacity, lysozymes, and IgM) parameters were substantially improved in the challenged fish fed on PRO and/or TUR diets compared to the challenged non-fed fish. The lipid profile and malondialdehyde were substantially decreased in the challenged fish fed on PRO and/or TUR diets compared to the challenged non-fed group. Notably, a down-turning of nuclear factor kappa B (NFκB) and tumor necrosis factor-alpha (TNF-α) expression with up-turning of nuclear factor erythroid 2-related factor 2 (Nrf2) and transforming growth factor-beta (TGF-β) expression was noticed in the challenged fish fed on PRO and/or TUR diets compared to the challenged non-fed fish. Noteworthy, dietary PRO and/or TUR improved the fish survival during E. tarda challenge. The mixture of PRO and TUR can be added to Nile tilapia diets to enhance their growth, immune response, and resistance to E. tarda. These outcomes help in the sustainable development of the Nile tilapia culture industry. Graphical Abstract

Toxicity assessment following conventional radiation therapy and pulsed low dose rate radiation therapy: an in vivo animal study

Abstract Background Pulsed low dose rate radiotherapy (PLDR) is a new radiation delivery method, in which the fractional dose is divided into sub-fractional doses with periodical time breaks in between. The goal of our study is to assess the toxicity on healthy tissues resulting from PLDR as compared to conventional radiotherapy (CRT) using the same physical X-ray dose. Methods We analyzed the weight and survival time for CRT and PLDR groups and studied the inflammatory cytokine transforming Growth Factor-β (TGF-β), usually released following irradiation. Histopathological and immunohistochemical analyses were conducted for intestinal and bone marrow tissues from rats subjected to 8 Gy whole- body irradiation using CRT and PLDR techniques. We investigated genotoxicity by performing a comet assay (CA) in splenic tissues. Results Our findings showed an improvement in survival time with PLDR versus CRT by 82%.The mean survival time for CRT rats’ group was 6.3 days, while it was 35.9 days for PLDR group.The weight of CRT group decreased gradually by 3.7%, while weight of PLDR group increased gradually by 2.4%.CRT resulted in more cellular atrophy in bone marrow and intestinal tissues than in PLDR treatments as shown by hematoxylin and eosin staining analysis. In addition, the transforming growth factor-β (TGF-β) expression in bone marrow and intestinal tissues of CRT was higher than those expressed in tissues from PLDR as demonstrated by the Immuno reactive score (IRS). It was10(0.53) and 9.8(0.55) for BM and intestinal tissues, respectively from CRT group and 5.8(0.63) for PLDR for both tissues. The measured CA parameters were larger with CRT compared to PLDR, where the Tail Length (TL), Tail DNA % (TD%) and Tail Moment (TM) measurements were 25.4(3.4), 56.5(7.6) % and 20.5(3.5) for CRT, 7.3(1.9), 30.0(7.2) % and 5.7(1.8) for PLDR, with P value 0.000064, 0.0004 and 0.00017, respectively. Conclusion This study indicates that PLDR can reduce the toxicity on normal tissues compared to CRT.

IL-13Rα2 Is Involved in Resistance to Doxorubicin and Survival of Osteosarcoma Patients

Background/Objectives: Interleukin 13 receptor alpha 2 (IL-13Rα2) is a receptor with a high affinity for IL-13 and is involved in the progression of human cancers. However, studies on the role of IL-13Rα2 in osteosarcoma are limited. Therefore, this study aimed to investigate the expression and roles of IL-13Rα2 in the progression of osteosarcoma. Methods: This study evaluated the roles of IL-13Rα2 in osteosarcomas by evaluating tumor tissues from 37 human osteosarcomas and osteosarcoma cells. Results: Immunohistochemical positivity of IL-13Rα2 was an independent indicator of shorter overall survival and relapse-free survival of 37 osteosarcoma patients and 26 subpopulations of patients who received adjuvant chemotherapy with multivariate analysis. In U2OS and KHOS/NP osteosarcoma cells, overexpression of IL-13Rα2 significantly increased proliferation, migration, and invasion of cells, all of which decreased with knockdown of IL-13Rα2. Overexpression of IL-13Rα2 increased expression of TGF-β, snail, cyclin D1, and BCL2 but decreased BAX, and knockdown of IL-13Rα2 caused a decrease in expression of these molecules. In addition, both in vitro and in vivo, proliferation of osteosarcoma cells increased, and apoptosis decreased with overexpression of IL-13Rα2 under treatment with doxorubicin. Knockdown of IL-13Rα2 sensitized osteosarcoma cells to the cytotoxic effect of doxorubicin. Conclusions: The results of this study suggest that the expression of IL13Rα2 might be used as a potential prognostic indicator in osteosarcoma patients. Furthermore, it is observed that IL13Rα2 influences the resistance to the chemotherapeutic agent doxorubicin. Therefore, a therapeutic trial targeting IL13Rα2 might be a new therapeutic strategy for osteosarcoma, especially those highly expressing IL13Rα2.

Sex differences and immune correlates of Long Covid development, symptom persistence, and resolution

Sex differences have been observed in acute coronavirus disease 2019 (COVID-19) and Long Covid (LC) outcomes, with greater disease severity and mortality during acute infection in males and greater proportions of females developing LC. We hypothesized that sex-specific immune dysregulation contributes to LC pathogenesis. To investigate the immunologic underpinnings of LC development and symptom persistence, we performed multiomic analyses on blood samples obtained during acute severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and 3 and 12 months after infection in a cohort of 45 participants who either developed LC or recovered. Several sex-specific immune pathways were associated with LC. Males who would later develop LC exhibited increases in transforming growth factor–β (TGF-β) signaling during acute infection, whereas females who would go on to develop LC had reduced TGFB1 expression. Females who developed LC demonstrated increased expression of XIST , an RNA gene implicated in autoimmunity, during acute infection compared with females who recovered. Many immune features of LC were also conserved across sexes, such as alterations in monocyte phenotype and activation state. Nuclear factor κB (NF-κB) transcription factors were up-regulated in many cell types at acute and convalescent time points. Those with ongoing LC demonstrated reduced ETS1 expression across lymphocyte subsets and elevated intracellular IL-4 in T cell subsets, suggesting that ETS1 alterations may drive aberrantly elevated T helper cell 2–like responses in LC. Altogether, this study describes multiple innate and adaptive immune correlates of LC, some of which differ by sex, and offers insights toward the pursuit of tailored therapeutics.

Abstract 4134695: The Effect and Mechanism of Histone Lactylation In Pathological Cardiac Hypertrophy Induced by Pressure Overload

Introduction: Cardiac hypertrophy is characterized by significant metabolic changes, notably an increase in glycolysis. Histone lactylation, a post-translational modification influenced by the glycolytic state of cells, plays a crucial role in regulating gene transcription. However, the relationship between histone lactylation and cardiac hypertrophy remains unclear. Research Questions: This study aims to elucidate the effects and underlying mechanisms of histone lactylation in the progression of cardiac hypertrophy induced by pressure overload. Methods and Results: We observed an increase in histone lactylation levels in hypertrophic hearts from both humans and mice. To investigate further, male mice underwent transverse aortic constriction (TAC) to induce cardiac hypertrophy, followed by either an intraperitoneal injection of Oxamate (an LDHA inhibitor) to reduce histone lactylation or sodium lactate to increase it. Reducing histone lactylation protected the hearts from TAC-induced hypertrophy and fibrosis, preserving cardiac function, while increased histone lactylation exacerbated cardiac hypertrophy and fibrosis, impairing cardiac function. Cardiomyocyte-specific deletion of LDHA also led to a reduction in cardiac hypertrophy. In vitro experiments showed that inhibiting histone lactylation reduced the expression of hypertrophic markers and the hypertrophic growth of cardiomyocytes stimulated by phenylephrine. Using co-immunoprecipitation, we confirmed that P300 and GCN5 are the transferases mediating histone lactylation in cardiomyocytes. Mechanistic studies using the CUT&Tag technique revealed lactate-dependent histone modification was enriched at the promoter of TGFB2, activating its transcription. Inhibiting cardiac-specific TGFB2 expression through adeno-associated viral delivery of shRNA reduced TAC-induced cardiac hypertrophy. Furthermore, TGFB2 overexpression induced a hypertrophic phenotype in cardiomyocytes and activated the PI3K/AKT/mTOR pathways. This hypertrophic phenotype, induced by TGFB2 overexpression, was suppressed by inhibitors of PI3K or AKT. Conclusion: Our findings reveal a critical role for histone lactylation in the development of pathological cardiac hypertrophy by regulating TGFB2 expression and the PI3K/AKT/mTOR pathway.

Abstract 4116300: Valvular Abnormalities and Risk of Abdominal Aortic Aneurysm Associated with Fluoroquinolone Use; A Retrospective Cohort Study

Introduction: Fluoroquinolones (FQ) are efficacious antibiotics whose antimicrobial effect is achieved through bacterial DNA gyrase inhibition. However, there is evidence suggesting that FQ may cause impaired collagen synthesis and an increase in tissue collagen degradation. Collagen serves as a predominant structural constituent of the aortic wall and heart valves. These structures undergo tissue maintenance, which necessitates the regulated expression of MMPs and TGF-β. In this study, we hypothesized that prolonged usage of FQs may increase risk of valvular abnormalities and abdominal aortic aneurysm. Methods: We conducted a retrospective cohort study utilizing de-identified patient data from the TriNetX platform, an extensive electronic health records (EHR) system. The cohorts consisted of smoker patients who used FQs for more than two consecutive weeks, Cohort A, N= 352,215 and smoker patients without a history of FQ use, Cohort B, N=350,912. Propensity score matching balanced baseline characteristics between cohorts: age, sex, ethnicity, and comorbidities. Results: The risk of developing AAA among smokers with a history of FQ use was 1.222% while the risk was 0.806% among smokers without history of FQ use (P< 0.0001, 95% CI (0.63,0.691), Risk Ratio (RR)= 0.66). The risk of developing nonrheumatic (NR) aortic insufficiency among smokers with history of FQ use was 1.479% while smokers with no history of fluoroquinolone use had a 0.95% risk (P<<0.0001, 95% CI (0.616,0.67), RR= 0.643). The risk of developing NR mitral insufficiency in smokers with history of FQ use was 3.873% versus their smoker FQ naive counterparts risk of 2.317% (P< 0.0001, 95% CI (0.582,0.615), RR= 0.598). NR tricuspid insufficiency in smokers with FQ usage was also increased versus their smoker FQ naive counterparts at 3.089% and 1.758%, respectively (P< 0.0001, 95% CI (0.552,0.587), RR =0.569). Conclusions: This study uncovered a significant association between FQ use, risk of AAA, and NR-valvular insufficiencies among smokers. In particular, male smokers are at highest risk of developing AAA. This same population is at risk for prostatitis that would likely need treatment with FQs, further increasing their risk. Despite the efficacy of this class of antibiotics, caution is paramount when prescribing FQs in individuals who are actively smoking or have any connective tissue pathologies.

Abstract 4113617: Single-cell Analysis Reveals Endothelial Cell CD45 Deficiency Prevents Endothelial-to-Mesenchymal Transition (EndMT) in Atherosclerosis

Introduction: Protein-tyrosine-phosphatase CD45 is exclusively expressed in all nucleated cells of the hematopoietic system but is rarely expressed in endothelial cells (ECs). However, a recent study indicated that activation of the endogenous CD45 promoter in human endothelial colony forming cells induced expression of EndMT marker genes. Also, the CD45 phosphatase inhibitor blocked EndMT activities in TGFβ1-treated ovine mitral valve endothelial cells. EndMT contributes to atherosclerotic pathobiology and is associated with more complex plaques. We identified CD45-positive ECs undergoing EndMT in atherosclerotic ApoE-deficient (ApoE -/- ) mice. Here, we aim to investigate whether endothelial CD45-specific deletion can prevent EndMT in a mouse model of atherosclerosis. Methods and Results: We generated a tamoxifen-inducible EC-specific CD45-deficient mouse strain (EC-iCD45KO) on an ApoE -/- background (C57BL/6) and fed these mice a Western diet for 16 weeks. We isolated and enriched mouse aortic endothelial cells with CD31 beads to perform single-cell RNA-seq. Cells populated 15 major clusters after integrating the single-cell transcriptomic profiles. In the EC-iCD45KO group, the population of endothelial, resident macrophage, and mesenchymal-endothelial transition cells increased while vascular smooth muscle, EndMT, and fibroblast cells decreased. In EndMT cells, EC markers (Cdh5, Cldn5, Pecam1) increased, but SMC markers (Acta2) and EndMT markers (Col1a2 and Col3a1）decreased. In addition, we characterized EndMT cells into two sub-clusters: 1 and 2. EC-specific CD45 deletions reduced the cell numbers in the EndMT2 sub-cluster in atherosclerotic mice but had less impact on the EndMT1 sub-cluster. FGFR pathways were potentiated while TGFβ1 and Tgfbr2 decreased in EndMT1 sub-cluster， and the genes controlling endothelial cell development were upregulated but the genes controlling actin cytoskeleton organization were downregulated in both EndMT1 and 2 sub-clusters. Conclusion: Our single-cell analysis indicates that the loss of endothelial CD45 protects against EndMT-driven atherosclerosis, inhibiting TGFβ and EndoMT marker expression while stimulating FGFR pathways. Consistently, EC-iCD45KO/ApoE -/- mice showed reduced plaque macrophages, expression of cell adhesion molecules, foam cell formation, and lesion development when compared to ApoE -/- controls. Thus, targeting endothelial CD45 may be a novel therapeutic strategy for EndMT and atherosclerosis.

Abstract 4134967: Pericardial Adipose Tissue Hypertrophy Contributes to Heart Failure via β3 Adrenergic Signaling-Mediated Activation of the TGFβ1 Pathway

Background: The heart is surrounded by pericardial adipose tissue (PeAT). An increase in PeAT volume has been associated with the development of heart failure; however, the precise molecular mechanisms by which hypertrophic PeAT contributes to heart failure remain to be elucidated. Aims: To clarify the involvement of PeAT hypertrophy in the development of heart failure. Methods: Male C57BL/6 mice (8-10 weeks old) were fed a high-fat diet (HFD) for 8 weeks to induce PeAT hypertrophy. Cardiac pressure overload was induced by transverse aortic constriction (TAC) with a 26-gauge needle. Results: PeAT volume significantly increased in HFD-fed mice compared to those on a normal diet (ND) (25.28 ± 4.1 mg vs. 9.35 ± 0.73 mg, p<0.01). Eight weeks after TAC, HFD-fed mice (TAC+HFD) exhibited more pronounced cardiac dysfunction (fractional shortening: 45.21 ± 2.21% vs. 55.99 ± 1.91%, p=0.002, left ventricular mass: 130.3 ± 7.80% vs. 106.8 ± 3.77%, p=0.0154), upregulation of cardiac fibrosis-related genes ( Col1a1 , Col3a1 ), and greater fibrotic area in the left ventricle compared to ND-fed TAC mice (TAC+ND). Simultaneously, the TGFβ/Smad pathway was activated in the left ventricles of TAC+HFD mice. Notably, surgical removal of hypertrophic PeAT improved cardiac function in TAC+HFD mice, while transplantation of PeAT from HFD-fed mice into TAC+ND mice worsened cardiac function. In TAC+HFD mice, local noradrenaline concentration in PeAT decreased, along with reduced activation of hormone-sensitive lipase, a downstream target of β3 adrenergic receptor signaling. Local free fatty acid concentrations also decreased in the PeAT of HFD-fed mice. In the RAW 264.7 macrophage cell line, a decrease in palmitic acid, a major fatty acid, led to increased expression of TGFβ1. Conclusions: These results suggest that HFD-induced PeAT hypertrophy suppresses β3 adrenergic receptor signaling and free fatty acid release, increasing TGFβ1 expression in the macrophages and resulting in cardiac fibrosis and dysfunction. Our findings provide new insights into the pathogenesis of PeAT hypertrophy in heart failure development.

Abstract 4145876: Prognostic significance of somatic mutations in myeloid cells of men with chronic heart failure – interaction between loss of Y chromosome and clonal hematopoiesis

Introduction: Age-associated clonal hematopoiesis of indeterminate potential (CHIP) has been associated with increased incidence and worse prognosis of chronic heart failure. CHIP is driven by somatic mutations in hematopoietic stem and progenitor cells (HSPC). Mosaic loss of the Y chromosome (LOY), the most common somatic mutation in blood cells of men, also correlates with clonal expansion of myeloid cells, increases with age and was experimentally shown to lead to diffuse cardiac fibrosis and subsequent heart failure in mice. However, the prognostic significance of LOY as well as its potential interaction with CHIP in patients with chronic heart failure is unknown. Aims/Methods: We investigated the prevalence and prognostic significance of the extent of LOY and the two most common CHIP-driver mutations DNMT3A and TET2 in 705 male patients with established chronic heart failure across the entire spectrum of left ventricular ejection fraction. Results: Both, LOY and DNMT3A/TET2 mutations, increased with age, and LOY co-occurred with DNMT3A/TET2 mutations in 27.1% of men at age > 70 years. LOY was an independent predictor of death during 3-years of follow-up across the entire spectrum of left ventricular ejection fraction. The co-occurrence of harboring LOY and DNMT3A/TET2 mutations significantly contributed to the observed increased mortality observed in carriers of DNMT3A/TET2 mutations. The detrimental effect of LOY on prognosis was confirmed in a validation cohort of patients with ischemic heart disease.scRNA sequencing of peripheral blood cells in patients with chronic ischemic heart failure showed increased profibrotic signaling in LOY monocytes with elevated markers of monocyte mediated inflammation and profibrotic cardiac remodeling (S100A8, TLR2, CLEC4D) and reduced expression of TGF-b inhibiting genes (SMAD7, TGIF2). The proinflammatory phenotype of LOY monocytes was further amplified in LOY monocytes of patients simultaneously harboring DNMT3A mutations, who displayed heightened expression of alarmins (S100A8, HMGB2) and interferon signaling related genes (IFNGR1, TRIM56, CD84) compared to patients without CHIP mutations. Conclusion: The age-associated acquisition of somatic mutations in blood cells of men with chronic heart failure is associated with increased mortality, with loss of Y chromosome emerging as an independent predictor of all-cause death across the entire spectrum of left ventricular function.

The Significance of TGF-β Expression in Predicting Lymphovascular Invasion and Lymph Node Metastasis in Colorectal Cancer

Background: Colorectal cancer (CRC) is a major health burden globally. The prognosis of CRC is strongly influenced by the presence of lymphovascular invasion (LVI) and lymph node (LN) metastasis. Transforming growth factor-beta (TGF-β) is a cytokine with a complex role in CRC progression. This study aimed to evaluate the significance of TGF-β expression in predicting LVI and LN metastasis in CRC. Methods: This cross-sectional study involved 50 patients diagnosed with CRC. The expression of TGF-β was assessed using immunohistochemical staining and the Allred scoring system. The relationship between TGF-β expression and the presence of LVI and LN metastasis was analyzed using the Chi-square test. Results: High TGF-β expression was significantly associated with both LVI (p = 0.011) and LN metastasis (p = 0.012) in CRC. Patients with high TGF-β expression had a higher risk of LVI and LN metastasis compared to those with low TGF-β expression. Conclusion: TGF-β expression is a significant predictor of LVI and LN metastasis in CRC. This finding has potential implications for risk stratification and treatment decisions in CRC patients.

Gene Expression Modulation of Markers Involved in Bone Formation and Resorption by Bisphenol A, Bisphenol F, Bisphenol S, and Bisphenol AF

Background: Bisphenol A (BPA) and its analogs (BPF, BPS, and BPAF) are recognized for inducing detrimental effects on various tissues, including bone. Objectives: The aim of this study is to investigate their impact on information and repair processes, specifically focusing on vascular endothelial growth factor (VEGF), transforming growth factor β1 (TGF-β1), and the receptors for transforming growth factor β (TGFR1, TGFR2, and TGFR3). Methods: Human osteoblasts isolated through primary culture from bone samples of healthy volunteers were subjected to cultivation in the presence of various dosage levels (10−5, 10−6, or 10−7 M) of BPA, BPF, BPS, or BPAF for 24 h. Gene expressions of RANKL, OPG, TGF-β1, TGFR1, TGFR2, TGFR3, and VEGF were analyzed by real-time polymerase chain reaction (RT-PCR). All experiments included untreated cells as controls. Results: Expressions of RANKL and OPG were dose-dependently downregulated by the presence of all tested bisphenols (BPs) except for BPAF, whose presence upregulated OPG expression at all three doses. TGF-β1 expression was downregulated by all BP treatments, and TGF-β1 receptor expression was also downregulated as a function of the BP and dose. VEGF expression was downregulated in the presence of BPF and BPAF at all three doses and in the presence of BPA at the two higher doses (10−5, and 10−6 M), but it was not changed by the presence of BPS at any dose. Conclusions: The inhibition of both RANKL and OPG by the BPs, with a higher %inhibition of RANKL than of OPG, appears to rule out BP-induced activation of osteoclastogenesis via RANKL/RANK/OPG. Nevertheless, the effect of the BPs on the expression by osteoblasts of TGF-β1, TGF-β receptors, and VEGF indicates that these compounds can be responsible for major molecular changes in this cell population, contributing to their adverse effects on bone tissue.

Expression of individual members of the TGF-β/SMAD signalling pathway in the progression and survival of patients with colorectal carcinoma

Current knowledge of tumor biology offers many “targets” for therapeutic intervention. The molecular basis of many processes that play a role in the pathogenesis of colorectal cancer has been identified. One part of colorectal cancer clinical trials is focused on testing substances in a group of patients with tumors in which the TGF-β signalling pathway is hyperactivated. The TGF-β/SMAD signalling pathway members are considered important markers; however, genetic, proteomic, or metabolomic analyses still yield controversial results. According to our results, TGF-βRII, and SMAD4 can be used in monitoring CRC progression. With increasing CRC stage, TGF-βRII expression decreases and SMAD4 expression increases. The patients with TGF-βRII expression lower than 700 pg/ml had a slightly lower survival time (28.103 months) than patients with higher TGF-βRII expression (31.620 months). Conversely, patients with SMAD4 expression lower than 200 pg/ml had a higher survival rate (30.979 months) than patients with higher expression (26.316 months). Regarding TGF-β1 expression, the patient´s survival assessment determined no significant difference between patients with high or low tissue TGF-β1 expression. A personalized approach and consideration of a wide range of factors are important when using these markers in treatment assessment.

Effect of moderate-intensity aerobic exercise on bladder TGF-β1 and type I collagen expressions in diabetic rat model

Transforming growth factor-beta 1 (TGF-β1) and type I collagen play crucial roles in the pathogenesis of diabetic bladder disease (DBD). Moderate-intensity aerobic exercise increases antioxidant activity to help manage DBD. The aim of this study was to evaluate the effect of moderate-intensity aerobic exercise on the expression of TGF-β1 and type I collagen in the detrusor and lamina propria of the bladder in a type 2 diabetes mellitus (T2DM) rat model. A true experimental design with a post-test-only control group design was conducted with white rats (Rattus norvegicus), divided into three groups: a T2DM model group sacrificed after T2DM induction and diagnosed with T2D from a fasting blood glucose (FBC) test (Group C), a T2DM model group that did not receive exercise (Group NE), and a T2DM model group that received moderate-intensity aerobic exercise (Group E). Moderate-intensity aerobic exercise was conducted over six weeks, with a frequency of five days per week for 60 minutes per session. The findings revealed a significant reduction in TGF-β1 expression in the lamina propria in Group E compared to Group C (p=0.004) Additionally, both Group E (p=0.002) and Group NE (p=0.028) showed a significant reduction in type I collagen expression in the lamina propria compared to Group C. These findings provide a basis for further investigation regarding the mechanism of non-pharmacologic DBD management by employing moderate-intensity exercise.

Thrombopoietin mimetic reduces mouse lung inflammation and fibrosis after radiation by attenuating activated endothelial phenotypes.

Radiation-induced lung injury (RILI) initiates radiation pneumonitis and progresses to fibrosis as the main side effect experienced by patients with lung cancer treated with radiotherapy. There is no effective drug for RILI. Sustained vascular activation is a major contributor to the establishment of chronic disease. Here, using a whole thoracic irradiation (WTI) mouse model, we investigated the mechanisms and effectiveness of thrombopoietin mimetic (TPOm) for preventing RILI. We demonstrated that administering TPOm 24 hours before irradiation decreased histologic lung injury score, apoptosis, vascular permeability, expression of proinflammatory cytokines, and neutrophil infiltration in the lungs of mice 2 weeks after WTI. We described the expression of c-MPL, a TPO receptor, in mouse primary pulmonary microvascular endothelial cells, showing that TPOm reduced endothelial cell-neutrophil adhesion by inhibiting ICAM-1 expression. Seven months after WTI, TPOm-treated lung exhibited less collagen deposition and expression of MMP-9, TIMP-1, IL-6, TGF-β, and p21. Moreover, TPOm improved lung vascular structure, lung density, and respiration rate, leading to a prolonged survival time after WTI. Single-cell RNA sequencing analysis of lungs 2 weeks after WTI revealed that TPOm shifted populations of capillary endothelial cells toward a less activated and more homeostatic phenotype. Taken together, TPOm is protective for RILI by inhibiting endothelial cell activation.

GLUT1 and prorenin receptor mediate differential regulation of TGF-β and CTGF in renal inner medullary collecting duct cells during high glucose conditions

BackgroundDuring diabetes, prorenin is highly produced by the renal collecting ducts. The binding of prorenin to (pro)renin receptor (PRR) on the apical plasma membrane triggers intracellular profibrotic genes, including TGF-β and CTGF. However, the underlying mechanisms contributing to the stimulation of these pathways remain unclear. Hence, we hypothesize that the glucose transporter-1 (GLUT1) favors the PRR-dependent stimulation of TGF-β and CTGF in the distal nephron segments during high glucose (HG) conditions.MethodsTo test this hypothesis, primary cultured renal inner medullary collecting duct (IMCD) cells were treated with normal glucose (NG, 5 mM) or high glucose (HG, 25 mM) for 48 h in the presence or absence of the GLUT1-specific inhibitor BAY 876 (2 nM). Additionally, IMCD cells were treated with the PRR antagonist PRO20. The expression of TGF-β and CTGF was quantified by immunoblot and qRT-PCR.ResultsHG increased GLUT1 mRNA and protein abundance, while BAY 876 inhibited these responses. HG treatment upregulated PRR, but the concomitant treatment with BAY 876 partially prevented this effect. TGF-β and CTGF expressions were augmented in IMCD cells treated with HG. However, PRO20 prevented the increases in TGF-β but not those of CTGF. GLUT1 inhibition partially prevented the increases in reactive oxygen species (ROS) during HG while PRO20 did not. ROS scavenging impaired CTGF upregulation during HG conditions. Additionally, long-term exposure to HG increases lipid peroxidation and reduced cell viability.ConclusionsThe data indicate that glucose transportation via GLUT1 is implicated in the PRR-dependent upregulation of TGF-β while CTGF is mediated mainly via a mechanism depending on ROS formation in renal medullary collecting duct cells.

Network pharmacology combined with transcriptomics reveals that formononetin, a biologically component of Astragalus membranaceus (Fisch.) Bunge, inhibits the PI3K/AKT signaling pathway to improve chronic renal failure

Ethnopharmacological relevanceFormononetin (FMN), one of the main isoflavones isolated from Astragalus membranaceus (Fisch.) Bunge, has multiple pharmacological and renal-protective effects. Our previous study suggested FMN as a candidate compound for the treatment of chronic renal failure (CRF). However, the mechanism underlying the repressive effect of FMN on the development of CRF is still unknown. Aims of the studyTo investigate the protective effect of FMN on CRF using in vivo and in vitro models and elucidate the potential underlying mechanism. Materials and methodsAn in vivo model of adenine-induced CRF and an in vitro model of human proximal tubule epithelial cells (HK-2) stimulated with transforming growth factor (TGF)-β1 were used. Serum levels of renal function parameters and inflammatory cytokines were evaluated. Histological analysis was performed to determine the extent of renal injury and fibrosis. Network pharmacology and mRNA sequencing were used to explore the potential mechanism. PPI analysis and molecular docking were used to identify key targets. Polymerase chain reaction and western blotting were used to determine the mechanism underlying the effect of FMN on CRF. ResultsFMN decreased the levels of renal function biochemical markers, including serum creatinine, blood urea nitrogen, and 24 h urine protein content. Treatment with FMN improved renal tubule injury and extracellular matrix (ECM) components, including collagens I and III. In addition, FMN significantly inhibited epithelial-mesenchymal transition (EMT); decreased the expression of fibronectin, N-cadherin, vimentin, α-SMA, and TGF-β1; and restored the expression of E-cadherin. The effect of FMN on renal interstitial fibrosis contributed to decreasing the expression of PI3K, p-Akt, and interleukin (IL) 4, restoring the expression of nitric oxide synthase 3 (NOS3), and reducing the release of inflammatory cytokines (IL-1β, IL-6, and tumor necrosis factor-alpha), both in vivo and in vitro. FMN treatment improved renal function and deposition of ECM components, reduced protein levels of EMT markers in rat kidneys and HK-2 cells, decreased the release of inflammatory cytokines, and inhibited the PI3K/Akt signaling pathway. ConclusionsFMN treatment significantly reduced the release of inflammatory cytokines and inhibited the effects of the PI3K/Akt signaling pathway on the key targets IL-4 and NOS3. Our results suggest FMN therapy as a novel therapeutic strategy for treating CRF.

Systems Pharmacology to Explore the Potential Mechanism of Ginseng Against Heart Failure.

The aim of this study is to elucidate the pharmacological mechanism underlying the effects of Ginseng Radix et Rhizoma (ginseng) in heart failure (HF), providing a theoretical foundation for its clinical application. The potential mechanism of ginseng in the context of HF was investigated using systems pharmacology that combined network pharmacology, Gene Expression Omnibus (GEO) analysis, molecular docking, and experimental verification. Network pharmacology was employed to identify drug-disease targets. Core gene targets were subsequently subjected to enrichment analysis by integrating network pharmacology with GEO. Molecular docking was utilized to predict the binding affinities between identified targets and ginseng compounds. Furthermore, the therapeutic efficacy of ginseng was validated in an isoproterenol (ISO)-induced rat model of HF. The modulation of key signaling pathways by ginseng was confirmed through Western blot analysis. A total of 154 potential targets of ginseng in the treatment of HF were identified through network pharmacology analysis. The analysis of GSE71613 revealed that the PI3K-Akt pathway, reactive oxygen species, oxidative phosphorylation, MAPK signaling, and Ras signaling pathways are predominantly associated with patients with HF. By integrating the findings from network pharmacology and GEO analysis, ginsenoside Rg1 and ginsenoside Rb3 were identified as the potential components in ginseng, while FN1 and PRKAA2 were recognized as key targets involved in the PI3K-AKT and AMPK pathways, respectively. Molecular docking analysis revealed a strong affinity between the potential components and the identified core targets. In vivo experiments indicated that the extract of ginseng (EPG) significantly ameliorated ISO-induced cardiac dysfunction by improving cardiac parameters such as cardiac left ventricular internal systolic diameter, left ventricular end-diastolic volume, left ventricular end systolic volume, and left ventricular ejection fraction, while also reducing malondialdehyde production. In addition, EPG was found to enhance superoxide dismutase activity and ATP levels, while concurrently reducing the levels of interleukin (IL)-1β, IL-6, and TNF-α. The extract also reduced myocardial oxygen consumption, inflammatory cell infiltration, and the number of damaged myocardial fibers. Moreover, EPG was observed to upregulate the expression of p-PI3K, p-AKT, p-AMPK, and Bcl-2, while downregulating the expression of p-NFκB, TGF-β, and Bax. The therapeutic effects of ginseng on HF are primarily mediated through the PI3K-Akt and AMPK pathways. Ginsenoside Rg1 and ginsenoside Rb3 have been identified as potential therapeutic agents for HF.

Evidence of Browning and Inflammation Features in Visceral Adipose Tissue of Women with Endometriosis

Patients with endometriosis tend to have a low body mass index, suggesting an inverse relationship between body fat and risk of disease. This is supported by evidence that miRNAs differentially expressed in endometriosis induce browning of pre-adipocytes in vitro. Thus, we hypothesize that endometriosis may underlie adipose tissue (AT) dysfunction and browning. Identify inflammation and browning processes in AT collected from endometriosis patients. Visceral and subcutaneous AT samples were obtained during endometriosis (n = 32) or uterine myoma (n = 14; controls) surgery. Blood catecholamines were determined by high-performance liquid chromatography while IL-6 and TGF-β levels were quantified by ELISA. Adipocyte cross-sectional areas were analyzed in H&E-stained sections by computer-assisted morphometry. Macrophages (F4/80; Galectin-3) and browning activation (UCP-1; PGC-1α) in tissues were identified by dual label immunofluorescence. Expression of inflammatory (IL-6; MCP-1; Galectin-3; CD206; TIMP1; TGF-β) and browning-related (UCP-1; PGC-1α; DIO2; CITED1; CIDEA; TMEM26; TBX1; PRDM16; PPAR-γ) molecules in AT were assessed by RT-PCR and Western blotting. Compared to controls, patients presented smaller adipocytes, especially in VAT, and lower norepinephrine levels. Serum IL-6, but not TGF-β, was increased in patients. UCP-1, PGC-1α, IL-6, and MCP-1 were upregulated in VAT from endometriosis women, which also evidenced a reduction of CD206, relative to controls. However, no differences were found in mRNA expression of IL-6, TIMP1, and TGF-β nor Galectin-3 protein levels. In SAT, protein expression remained unchanged between patients and controls. Our findings support an endometriosis' role as a pro-catabolic state along with local signals of VAT browning and inflammation.

Hemin attenuates bleomycin-induced lung fibrosis in mice by regulating the TGF-β1/MAPK and AMPK/SIRT1/PGC-1α/HO-1/NF-κB pathways.

The objective of this study was to investigate the protective effect and potential mechanism of action of hemin on bleomycin-induced pulmonary fibrosis in mice. Male C57BL/6 mice were randomly divided into control, bleomycin and bleomycin + hemin groups. Mice in the bleomycin and bleomycin + hemin groups were injected intratracheally with bleomycin to establish the pulmonary fibrosis model. The bleomycin + hemin group mice were injected intraperitoneally with hemin starting 7 days before modeling until the end of Day 21 after modeling. Pathological changes in lung tissue were assessed by HE and Masson staining. Malondialdehyde (MDA), superoxide dismutase (SOD) and catalase (CAT) levels were determined in lung tissue. Immunohistochemistry was performed to assess the expression of α-SMA and collagen I. The serum levels of IL-6 and TNF-α were measured via ELISA. Western blotting was used to determine the expression of TGF-β1, SIRT1, PGC-1α and HO-1 and the phosphorylation levels of p38, ERK1/2, JNK, AMPK and NF-κB p65 in lung tissue. Hemin significantly reduced lung indices, increased terminal body weight. It also significantly increased SOD and CAT activities; decreased MDA, IL-6 and TNF-α levels; reduced the levels of α-SMA and collagen I-positive cells; upregulated SIRT1, PGC-1α and HO-1 expression; promoted AMPK phosphorylation; and downregulated TGF-β1 expression and p38, ERK1/2, JNK and NF-κB p65 phosphorylation. Hemin might attenuate oxidative damage and inflammatory responses and reduces extracellular matrix deposition by regulating the expression and phosphorylation of proteins associated with the TGF-β1/MAPK and AMPK/SIRT1/PGC-1α/HO-1/NF-κB pathways, thereby alleviating bleomycin-induced pulmonary fibrosis.