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With the increasing incidence of kidney diseases, there is an urgent need to develop therapeutic strategies to combat post-injury fibrosis. Immune cells, including platelets, play a pivotal role in this repair process, primarily through their released cytokines. However, the specific role of platelets in kidney injury and subsequent repair remains underexplored. Here, the detrimental role of platelets in renal recovery following ischemia/reperfusion injury and its contribution to acute kidney injury  to chronic kidney disease transition is aimed to investigated. In this study, it is shown that depleting platelets accelerates injury resolution and significantly reduces fibrosis. Employing advanced single-cell and spatial transcriptomic techniques, macrophages as the primary mediators modulated by platelet signals is identified. A novel subset of macrophages, termed cycling M2, which exhibit an M2 phenotype combined with enhanced proliferative activity is uncovered. This subset emerges in the injured kidney during the resolution phase and is modulated by platelet-derived thrombospondin 1 (THBS1) signaling, acquiring profibrotic characteristics. Conversely, targeted inhibition of THBS1 markedly downregulates the cycling M2 macrophage, thereby mitigating fibrotic progression. Overall, this findings highlight the adverse role of platelet THBS1-boosted cycling M2 macrophages in renal injury repair and suggest platelet THBS1 as a promising therapeutic target for alleviating inflammation and kidney fibrosis.

Duchenne Muscular Dystrophy (DMD) is a progressive and fatal neuromuscular disease. Cycles of myofibre degeneration and regeneration are hallmarks of the disease where immune cells infiltrate to repair damaged skeletal muscle. Benfotiamine is a lipid soluble precursor to thiamine, shown clinically to reduce inflammation in diabetic related complications. We assessed whether benfotiamine administration could reduce inflammation related dystrophic pathology. Benfotiamine (10 mg/kg/day) was fed to male mdx mice (n = 7) for 15 weeks from 4 weeks of age. Treated mice had an increased growth weight (5-7 weeks) and myofibre size at treatment completion. Markers of dystrophic pathology (area of damaged necrotic tissue, central nuclei) were reduced in benfotiamine mdx quadriceps. Grip strength was increased and improved exercise capacity was found in mdx treated with benfotiamine for 12 weeks, before being placed into individual cages and allowed access to an exercise wheel for 3 weeks. Global gene expression profiling (RNAseq) in the gastrocnemius revealed benfotiamine regulated signalling pathways relevant to dystrophic pathology (Inflammatory Response, Myogenesis) and fibrotic gene markers (Col1a1, Col1a2, Col4a5, Col5a2, Col6a2, Col6a2, Col6a3, Lum) towards wildtype levels. In addition, we observed a reduction in gene expression of inflammatory gene markers in the quadriceps (Emr1, Cd163, Cd4, Cd8, Ifng). Overall, these data suggest that benfotiamine reduces dystrophic pathology by acting on inflammatory and fibrotic gene markers and signalling pathways. Given benfotiamine's excellent safety profile and current clinical use, it could be used in combination with glucocorticoids to treat DMD patients.

BACKGROUND: Aging-associated left ventricular dysfunction promotes cardiopulmonary fibrogenic remodeling, Group 2 pulmonary hypertension (PH), and right ventricular failure. At the time of diagnosis, cardiac function has declined, and cardiopulmonary fibrosis has often developed. Here, we sought to develop a molecular positron emission tomography (PET)-magnetic resonance imaging (MRI) protocol to detect both cardiopulmonary fibrosis and fibrotic disease activity in a left ventricular dysfunction model. METHODS AND RESULTS: Left ventricular dysfunction was induced by transverse aortic constriction (TAC) in 6-month-old senescence-accelerated prone mice, a subset of mice that received sham surgery. Three weeks after surgery, mice underwent simultaneous PET-MRI at 4.7 T. Collagen-targeted PET and fibrogenesis magnetic resonance (MR) probes were intravenously administered. PET signal was computed as myocardium- or lung-to-muscle ratio. Percent signal intensity increase and Δ lung-to-muscle ratio were computed from the pre-/postinjection magnetic resonance images. Elevated allysine in the heart (P=0.02) and lungs (P=0.17) of TAC mice corresponded to an increase in myocardial magnetic resonance imaging percent signal intensity increase (P

PurposeAnterior urethral stricture disease (aUSD) is a complex, heterogeneous condition that is idiopathic in origin for most men. This gap in knowledge rarely affects the current management strategy for aUSD, as urethroplasty does not generally consider etiology. However, as we transition towards personalized, minimally invasive treatments for aUSD and begin to consider aUSD prevention strategies, disease pathophysiology will become increasingly important. The purpose of this study was to perform a deep phenotype of men undergoing anterior urethroplasty for aUSD. We hypothesized that unique biologic signatures and potential targets for intervention would emerge based on stricture presence/absence, stricture etiology, and the presence/absence of stricture inflammation.Materials and methodsMen with aUSD undergoing urethroplasty were recruited from one of 5 participating centers. Enrollees provided urethral stricture tissue and blood/serum on the day of surgery and completed patient-reported outcome measure questionnaires both pre- and postoperatively. The initial study had 3 aims: (1) to determine pediatric and adult subacute and repeated perineal trauma (SRPT) exposures using a study-specific SRPT questionnaire, (2) to determine the degree of inflammation and fibrosis in aUSD and peri-aUSD (normal urethra) tissue, and (3) to determine levels of systemic inflammatory and fibrotic cytokines. Two controls groups provided serum (normal vasectomy patients) and urethral tissue (autopsy patients). Cohorts were based on the presence/absence of stricture, by presumed stricture etiology (idiopathic, traumatic/iatrogenic, lichen sclerosus [LS]), and by the presence/absence of stricture inflammation.ResultsOf 138 enrolled men (120 tissue/serum; 18 stricture tissue only), 78 had idiopathic strictures, 33 had trauma-related strictures, and 27 had LS-related strictures. BMI, stricture length, and stricture location significantly differed between cohorts (P < .001 for each). The highest BMIs and the longest strictures were observed in the LS cohort. SRPT exposures did not significantly differ between etiology cohorts, with > 60% of each reporting low/mild risk. Stricture inflammation significantly differed between cohorts, with mild to severe inflammation present in 27% of trauma-related strictures, 54% of idiopathic strictures, and 48% of LS strictures (P = .036). Stricture fibrosis did not significantly differ between cohorts (P = .7). Three serum cytokines were significantly higher in patients with strictures compared to stricture-free controls: interleukin-9 (IL-9; P = .001), platelet-derived growth factor-BB (P = .004), and CCL5 (P = .01). No differences were observed in the levels of these cytokines based on stricture etiology. However, IL-9 levels were significantly higher in patients with inflamed strictures than in patients with strictures lacking inflammation (P = .019). Degree of stricture inflammation positively correlated with serum levels of IL-9 (Spearman's rho 0.224, P = .014).ConclusionsThe most common aUSD etiology is idiopathic. Though convention has implicated SRPT as causative for idiopathic strictures, here we found that patients with idiopathic strictures had low SRPT rates that were similar to rates in patients with a known stricture etiology. Stricture and stricture-adjacent inflammation in idiopathic stricture were similar to LS strictures, suggesting shared pathophysiologic mechanisms. IL-9, platelet-derived growth factor-BB, and CCL5, which were elevated in patients with strictures, have been implicated in fibrotic conditions elsewhere in the body. Further work will be required to determine if this shared biologic signature represents a potential mechanism for an aUSD predisposition.

Aberration of the "gut-liver axis" contributes to the development and progression of metabolic dysfunction-associated steatotic liver disease (MASLD). Here, we use multi-omics to analyze the gut microbiota composition and metabolic profile of patients with type-2 diabetes mellitus (T2DM). T2DM patients were screened for liver disease by blood tests, ultrasound, and liver stiffness measurements. Stool microbiota was analyzed by 16S rRNA gene sequencing; metabolomic profiling by Nuclear Magnetic Resonance spectroscopy and Ultra-High Performance-Mass Spectrometry. Microbiome and metabolic signatures were analyzed in the whole cohort and in matched subsets to identify signatures specific for steatosis (MASLD±) or fibrosis (Fibrosis±). Gut permeability was assessed in-vitro using monolayers of MDCK cells and trans-epithelial electric resistance (TEER). Cytokine profile was assessed in serum and stools.Overall, 285 patients were enrolled: 255 serum, 252 urine and 97 stool samples were analyzed. Anaeroplasma and Escherichia/Shigella ASVs were higher, while Butyricicoccus ASVs were lower in those with normal liver. In MASLD±, Butyricicoccus ASV was significantly higher in those with steatosis. In the Fibrosis±, Butyricicoccus ASV was significantly lower in those with fibrosis. Glycochenodeoxycholic acid-3-sulfate (G-UDCA-3S) appeared to be higher in MASLD with fibrosis. Fecal water from patients with MASLD and fibrosis caused the greatest drop in the TEER vs those with normal liver; this was reversed with protease inhibitors. Finally, fecal IL-13 was lower in MASLD with fibrosis. We identified microbiome signatures which were specific for steatosis and fibrosis and independent of other metabolic risk factors. Moreover, we conclude that protease-related gut permeability plays a role in those MASLD patients with fibrosis, and that disease progression is linked to a gut-liver axis which is at least partially independent of T2DM.

Aging coincides with the progressive loss of muscle mass and strength, increased adiposity, and diminished physical function. Accordingly, interventions aimed at improving muscle, metabolic, and/or physical health are of interest to mitigate the adverse effects of aging. In this study, we tested a stem cell secretome product, which contains extracellular vesicles and growth, cytoskeletal remodeling, and immunomodulatory factors. We examined the effects of 4 weeks of 2×/week unilateral intramuscular secretome injections (quadriceps) in ambulatory aged male C57BL/6 mice (22-24 months) compared to saline-injected aged-matched controls. Secretome delivery substantially increased whole-body lean mass and decreased fat mass, corresponding to higher myofiber cross-sectional area and smaller adipocyte size, respectively. Secretome-treated mice also had greater whole-body physical function (grip strength and rotarod performance) and had higher energy expenditure and physical activity levels compared to control mice. Furthermore, secretome-treated mice had greater skeletal muscle Pax7+ cell abundance, capillary density, collagen IV turnover, reduced intramuscular lipids, and greater Akt and hormone sensitive lipase phosphorylation in adipose tissue. Finally, secretome treatment in vitro directly enhanced muscle cell growth and IL-6 production, and in adipocytes, it reduced lipid content and improved insulin sensitivity. Moreover, indirect treatment with secretome-treated myotube culture media also enhanced muscle cell growth and adipocyte size reduction. Together, these data suggest that intramuscular treatment with a stem cell secretome improves whole-body metabolism, physical function, and remodels skeletal muscle and adipose tissue in aged mice.

BACKGROUND: Loss of AZGP1 expression is a biomarker associated with progression to castration resistance, development of metastasis, and poor disease-specific survival in prostate cancer. However, high expression of AZGP1 cells in prostate cancer has been reported to increase proliferation and invasion. The exact role of AZGP1 in prostate cancer progression remains elusive. METHOD: AZGP1 knockout and overexpressing prostate cancer cells were generated using a lentiviral system. The effects of AZGP1 under- or over-expression in prostate cancer cells were evaluated by in vitro cell proliferation, migration, and invasion assays. Heterozygous AZGP1± mice were obtained from European Mouse Mutant Archive (EMMA), and prostate tissues from homozygous knockout male mice were collected at 2, 6 and 10 months for histological analysis. In vivo xenografts generated from AZGP1 under- or over-expressing prostate cancer cells were used to determine the role of AZGP1 in prostate cancer tumor growth, and subsequent proteomics analysis was conducted to elucidate the mechanisms of AZGP1 action in prostate cancer progression. AZGP1 expression and microvessel density were measured in human prostate cancer samples on a tissue microarray of 215 independent patient samples. RESULT: Neither the knockout nor overexpression of AZGP1 exhibited significant effects on prostate cancer cell proliferation, clonal growth, migration, or invasion in vitro. The prostates of AZGP1-/- mice initially appeared to have grossly normal morphology; however, we observed fibrosis in the periglandular stroma and higher blood vessel density in the mouse prostate by 6 months. In PC3 and DU145 mouse xenografts, over-expression of AZGP1 did not affect tumor growth. Instead, these tumors displayed decreased microvessel density compared to xenografts derived from PC3 and DU145 control cells, suggesting that AZGP1 functions to inhibit angiogenesis in prostate cancer. Proteomics profiling further indicated that, compared to control xenografts, AZGP1 overexpressing PC3 xenografts are enriched with angiogenesis pathway proteins, including YWHAZ, EPHA2, SERPINE1, and PDCD6, MMP9, GPX1, HSPB1, COL18A1, RNH1, and ANXA1. In vitro functional studies show that AZGP1 inhibits human umbilical vein endothelial cell proliferation, migration, tubular formation and branching. Additionally, tumor microarray analysis shows that AZGP1 expression is negatively correlated with blood vessel density in human prostate cancer tissues. CONCLUSION: AZGP1 is a negative regulator of angiogenesis, such that loss of AZGP1 promotes angiogenesis in prostate cancer. AZGP1 likely exerts heterotypical effects on cells in the tumor microenvironment, such as stromal and endothelial cells. This study sheds light on the anti-angiogenic characteristics of AZGP1 in the prostate and provides a rationale to target AZGP1 to inhibit prostate cancer progression.

This study investigates Bacillus Calmette-Guérin (BCG) as a potential treatment for hepatocellular carcinoma (HCC), a condition often associated with unfavorable treatment outcomes. Exploiting BCGs recognized immune-boosting properties, preclinical trials are conducted using HCC mice, with a single subcutaneous dose of BCG administered post-tumor formation. Results indicate that BCG treatment effectively diminishes tumor burden and extends survival in both male and female HCC mice. Positive influences on hepatic fibrosis and metabolism are observed, leading to a reduction in lipid levels. Spatial analysis underscores BCGs tumor-specific effects, inducing the enrichment of metabolic pathways and inhibiting various cancer-related pathways. Furthermore, BCG promotes immune cell infiltration, including CD4+, CD8+ T cells, and M1 macrophages, in both v-akt murine thymoma viral oncogene homolog 1(AKT)/neutoblastoma RAS viral oncogene homolog (RAS) and β-catenin positive HCC models. Interestingly, blocking T cells, trained immunity, and Interferon-γ (IFN-γ) function reverses BCGs anti-HCC effects. In conclusion, BCG emerges as a promising treatment option for HCC, characterized by a favorable safety profile and efficacy in inhibiting fibrosis, improving metabolism, and engaging both trained immunity and T cells in therapeutic mechanisms.

BACKGROUND: As the prevalence of metabolic dysfunction-associated steatotic liver disease increases, it is imperative to have noninvasive alternatives to liver biopsy. Velacur offers a non-invasive, point-of-care ultrasound-based method for the assessment of liver stiffness and attenuation. The aim of this study was to perform a head-to-head comparison of liver stiffness and liver fat determined by Velacur and FibroScan using MRI-based measurements as the reference standard. METHODS: This prospective cross-sectional study included 164 adult participants with well-characterized metabolic dysfunction-associated steatotic liver disease. Patients underwent a research exam including Velacur, FibroScan and contemporaneous magnetic resonance elastography, and magnetic resonance imaging proton density fat fraction (MRI-PDFF) scans. The primary outcome was the presence of advanced fibrosis (>F2) as measured by magnetic resonance elastography and the presence of liver fat (>5%) as measured by MRI-PDFF. RESULTS: The mean age and body mass index were 57±12 years and 30.6±4.8 kg/m2, respectively. The mean liver stiffness on magnetic resonance elastography was 3.22±1.39 kPa and the mean liver fat on MRI-PDFF was 14.2±8%. The liver stiffness assessments by Velacur and FibroScan were similar for the detection of advanced fibrosis (AUC 0.95 vs. 0.97) and were not statistically different (p=0.43). Velacur was significantly better than FibroScan (AUC 0.94 vs. 0.79, p=0.01), for the detection of MRI-PDFF >5% (diagnosis of metabolic dysfunction-associated liver disease). CONCLUSIONS: Velacur was superior to FibroScan for liver fat detection with MRI-PDFF as the reference. Velacur and FibroScan were not statistically different for liver stiffness assessment as defined by magnetic resonance elastography.