Dexrazoxane Alleviated Doxorubicin-Induced Nephropathy in Rats

<b><i>Introduction:</i></b> Doxorubicin (DOX), an anthracycline antitumor agent, has been widely used against various solid tumors and hematological malignancies. However, the clinical application of DOX is restricted by its multiple organ toxicity including nephrotoxicity. This study investigated the protective effects and mechanisms of dexrazoxane (DZR) against DOX-induced nephropathy in rats. <b><i>Methods:</i></b> Male Sprague Dawley rats received 2.5 mg/kg DOX once a week for 5 consecutive weeks. 24-h urinary protein and renal function injury biomarkers were determined to evaluate the renal function. Histopathological changes and glomerulosclerosis were examined by hematoxylin and eosin and periodic acid-Schiff staining. The change of renal ultrastructure in the DOX-induced rats was observed by the electron microscopy. The renal apoptosis was detected by TUNEL staining and measured the protein expression of Caspase-3, Bcl-2, and Bax. Renal interstitial fibrosis was determined by Masson staining and immunohistochemistry examination. The levels of vimentin, alpha-smooth muscle actin (α-SMA), and transforming growth factor β (TGF-β) in kidney tissue were detected by Western blot. <b><i>Results:</i></b> DZR pretreatment markedly raised the survival rate and improved the renal dysfunction in DOX-treated rats. DZR ameliorated DOX-induced histopathological lesion of glomerular and tubular and apoptosis. DZR restored the oxidant/antioxidant balance via regulating the levels of MDA, SOD, and TAC. DZR reduced DOX-induced collagen IV deposition and renal interstitial fibrosis and downregulated the fibrosis-related protein expressions of vimentin, α-SMA, and TGF-β1. <b><i>Conclusion:</i></b> Our results suggest DZR exerted its protective effects against DOX-induced nephropathy through inhibition of lipid peroxidation, apoptosis, and fibrosis.

Histological and Immunohistochemical Characterization of Vascular Alterations in Meninges of Cats Infected with Gurltia paralysans

Gurltia paralysans, a metastrongyloid nematode, parasitizes in meningeal vessels in the thoracolumbar spinal cord of cats in South America and causes progressive paraparesis. Recently, the first report outside of South America described gurltiosis in a cat in Spain. As this parasitic disease has so far been largely neglected, especially outside of South America, the aim of the present case study was to add knowledge to the histologic and immunohistochemical characterization of central nervous lesions. To this purpose, formalin-fixed and paraffin-embedded (FFPE) tissue samples from the spinal cord and brain of five cats affected by clinical signs caused by Gurltia paralysans and of three control cats without CNS lesions were histopathologically examined using hematoxylin and eosin stain (HE), Elastica van Gieson stain, as well as periodic acid–Schiff (PAS) reaction. Moreover, immuno- histochemistry for alpha smooth muscle actin and Factor VIII-related antigen were performed to characterize vascular lesions. Lesions were consistent with previous descriptions and were mainly located in the spinal cord and consisted of chronic suppurative or lymphoplasmahistiocytic meningi tis as well as suppurative vasculitis, congestion and varicosis of meningeal veins. In view of the recent detection of this parasite in Europe and the increasing inner-European transport of rescued domestic cats, veterinarians in Europe should be aware of the clinical and pathomorphological presentation of this disease.

Genes that Escape X Chromosome Inactivation Modulate Sex Differences in Valve Myofibroblasts

Background: Aortic valve stenosis (AVS) is a sexually dimorphic disease, with women often presenting with sustained fibrosis and men with more extensive calcification. However, the intracellular molecular mechanisms that drive these clinically important sex differences remain under explored. Methods: Hydrogel scaffolds were designed to recapitulate key aspects of the valve tissue microenvironment and serve as a culture platform for sex-specific valvular interstitial cells (VICs; precursors to pro-fibrotic myofibroblasts). The hydrogel culture system was used to interrogate intracellular pathways involved in sex-dependent VIC-to-myofibroblast activation and deactivation. RNA-sequencing was used to define pathways involved in driving sex-dependent activation. Interventions using small molecule inhibitors and small interfering RNA (siRNA) transfections were performed to provide mechanistic insight into sex-specific cellular responses to microenvironmental cues, including matrix stiffness and exogenously delivered biochemical factors. Results: In both healthy porcine and human aortic valves, female leaflets had higher baseline activation of the myofibroblast marker, alpha-smooth muscle actin (α-SMA), compared to male leaflets. When isolated and cultured, female porcine and human VICs had higher levels of basal α-SMA stress fibers that further increased in response to the hydrogel matrix stiffness, both of which were higher than male VICs. A transcriptomic analysis of male and female porcine VICs revealed Rho-associated protein kinase (RhoA/ROCK) signaling as a potential driver of this sex-dependent myofibroblast activation. Further, we found that genes that escape X-chromosome inactivation, such as BMX and STS (encoding for Bmx non-receptor tyrosine kinase and steroid sulfatase, respectively) partially regulate the elevated female myofibroblast activation via RhoA/ROCK signaling. This finding was confirmed by treating male and female VICs with endothelin-1 and plasminogen activator inhibitor-1, factors that are secreted by endothelial cells and known to drive myofibroblast activation via RhoA/ROCK signaling. Conclusions: Together, in vivo and in vitro results confirm sex-dependencies in myofibroblast activation pathways and implicate genes that escape X-chromosome inactivation in regulating sex differences in myofibroblast activation and subsequent AVS progression. Our results underscore the importance of considering sex as a biological variable to understand the molecular mechanisms of AVS and help guide sex-based precision therapies.

Impediment of Cerebrospinal Fluid Drainage Through Glymphatic System in Glioma

BackgroundCerebrospinal fluid (CSF) plays an important role in maintaining tissue homeostasis in the central nervous system. In 2012, the new CSF outflow pathway, “the glymphatic system,” was discovered. The glymphatic system mediates CSF and interstitial fluid exchange through the perivascular pathway, which eliminates harmful solutes in the brain parenchyma. In recent studies, the importance of the glymphatic system has been demonstrated in healthy and neurodegenerative disease brains. However, there is limited research on the function of the CSF in brain tumors. Intracranial hypertension caused by glioma can affect CSF drainage, which impacts the delivery of chemotherapy drugs via intrathecal injection. This study focused on changes in the glymphatic system and the role of aquaporin 4 (AQP4) in glymphatic transport in glioma.MethodsIn glioma-bearing rats, the effect of tracer infusion on the intracranial pressure (ICP) was evaluated using an ICP microsensor. In vivo magnetic resonance imaging and ex vivo bright field were used to monitor CSF tracer distribution after cisterna magna injection. AQP4 expression was quantitatively detected, and AQP4 in the astrocytes around the vessels was observed using immunofluorescence.ResultsThe ICP of the tumor group was higher than that of the control group and the infusion rate of 2 µl/min did not affect ICP. In vivo and ex vivo imaging showed that the circulation of CSF tracers was significantly impaired in the tumor. High-power confocal microscopy revealed that, in the tumor, the surrounding of AQP4 by Evans Blue was decreased. In both tumor and contralateral areas, data indicated that the number of cluster designation 34 (CD34+) alpha-smooth muscle actin (α-SMA−) veins were more than that of CD34+α-SMA+ arteries. Moreover, in the tumor area, AQP4 in the astrocytes around the vessels was decreased.ConclusionsThese findings indicate that the para-arterial influx of subarachnoid CSF is limited in glioma, especially in those with reduced levels of the fundamental protein AQP4. Our results provide evidence toward a potential new treatment method for glioma in the future.

Benign mixed Müllerian (duct) vaginal tumor in a 12-y-old goat

In human and veterinary medicine, mixed Müllerian tumors (MMTs) are rarely diagnosed neoplasms of the tubular female genital tract. Although there are case reports of malignant MMTs in various species, benign MMTs have only been described once in a macaque. Here we present a case of benign MMT in a 12-y-old goat, and review the literature on uterine, cervical, and vaginal neoplasia in goats. The doe was presented with vaginal discharge and was euthanized because of the high suspicion of intraabdominal neoplasia. On gross examination, an ulcerated vaginal mass was identified. Histologically, 2 distinct cell populations were present: smooth muscle cells that were well differentiated and positive for alpha–smooth muscle actin, and ciliated columnar epithelial cells that lined ductal structures and had no signs of malignancy. These findings led to the diagnosis of neoplasia of Müllerian origin. Benign MMT should be considered as a differential diagnosis for uterine and vaginal neoplasms in goats.

Dermal Papilla Cell-Derived Extracellular Vesicles Increase Hair Inductive Gene Expression in Adipose Stem Cells via β-Catenin Activation

Recently, extracellular vesicle (EV)-mediated cell differentiation has gained attention in developmental biology due to genetic exchange between donor cells and recipient cells via transfer of mRNA and miRNA. EVs, also known as exosomes, play a role in maintaining paracrine cell communication and can induce cell proliferation and differentiation. However, it remains unclear whether adipose-derived stem cells (ASCs) can adopt dermal papilla (DP)-like properties with dermal papilla cell-derived extracellular vesicles (DPC-EVs). To understand the effect of DPC-EVs on cell differentiation, DPC-EVs were characterized and incubated with ASCs, of monolayer and spheroid cell cultures, in combination with the CAO1/2FP medium specialized for dermal papilla cells (DPCs). DPC-like properties in ASCs were initially evaluated by comparing several genes and proteins with those of DPCs via real-time PCR analysis and immunostaining, respectively. We also evaluated the presence of hair growth-related microRNAs (miRNAs), specifically mir-214-5P, mir-218-5p, and mir-195-5P. Here, we found that miRNA expression patterns varied in DPC-EVs from passage 4 (P4) or P5. In addition, DPC-EVs in combination with CAP1/2FP accelerated ASC proliferation at low concentrations and propagated hair inductive gene expression for versican (vcan), alpha-smooth muscle actin (α-sma), osteopontin (opn), and N-Cam (ncam). Comparison between the expression of hair inductive genes (vcan, α-sma, ctnb, and others), the protein VCAN, α-SMA and β-Catenin (CTNB), and hair inductive miRNAs (mir-214-5P, mir-218-5p, and mir-195-5p) of DPC-EVs revealed similarities between P4 DPC-EVs-treated ASCs and DPCs. We concluded that early passage DPC-EVs, in combination with CAP1/2FP, enabled ASCs to transdifferentiate into DPC-like cells.

EndMT: New findings on the origin of myofibroblasts in endometrial fibrosis of intrauterine adhesions

Background Intrauterine adhesion (IUA) is one of the leading causes of infertility and the main clinical challenge is the high recurrence rate. The key to solving this dilemma lies in elucidating the mechanisms of endometrial fibrosis. The aim of our team is to study the mechanism underlying intrauterine adhesion fibrosis and the origin of fibroblasts in the repair of endometrial fibrosis. Methods Our experimental study involving an animal model of intrauterine adhesion and detection of fibrosis-related molecules. The levels of molecular factors related to the endothelial-to-mesenchymal transition (EndMT) were examined in a rat model of intrauterine adhesion using immunofluorescence, immunohistochemistry, qPCR and Western blot analyses. Main outcome measures are levels of the endothelial marker CD31 and the mesenchymal markers alpha-smooth muscle actin (α-SMA) and vimentin. Results Immunofluorescence co-localization of CD31 and a-SMA showed that 14 days after moulding, double positive cells for CD31 and a-SMA could be clearly observed in the endometrium. Decreased CD31 levels and increased α-SMA and vimentin levels indicate that EndMT is involved in intrauterine adhesion fibrosis. Conclusions Endothelial cells promote the emergence of fibroblasts via the EndMT during the endometrial fibrosis of intrauterine adhesions.

Liver Fibrosis: Underlying Mechanisms and Innovative Therapeutic Approach. A Review Article

Liver fibrosis is considered: “a pathological repairing process in liver injuries leading to extracellular cell matrix (ECM) accumulation evidencing chronic liver diseases”. Chronic viral hepatitis, alcohol consumption, autoimmune diseases as well as non-alcoholic steatohepatitis are from the main causes of liver fibrosis (Lee et al., 2015; Mieli-Vergani et al., 2018). Hepatic stellate cells (HSCs) exist in the sinus space next to the hepatic epithelial cells as well as endothelial cells (Yin et al., 2013). Normally, HSCs are quiescent and mainly participate in fat storage and in the metabolism of vitamin A. HSCs are produced during liver injury and then transformed into myofibroblasts. The activated HSCs resulted in a sequence of events considered as marks fibrosis. The activation of HSCs mostly express alpha smooth muscle actin (α-SMA). Moreover, ECM is synthesized and secreted by HSCs that affects markedly the structure and function of the liver tissue leading to fibrosis (Tsuchida et al., 2017; Han et al., 2020). Hence, activated HSCs are attracting attention as potential targets in liver fibrosis. Many signaling molecules are involved in HSCs activation first and foremost, platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-β) (Tsuchida et al., 2017; Wang et al., 2020c) as interfering the PDGF or TGF-β signaling pathways is a growing field for liver fibrosis treatment.

Alpha Smooth Muscle Actin (αSMA) Immunohistochemistry Use in the Differentiation of Pancreatic Cancer from Chronic Pancreatitis

Aim: Fibrosis is observed both in pancreatic cancer (PDAC) and chronic pancreatitis (CP). The main cells involved in fibrosis are pancreatic stellate cells (PSCs), which activate alpha smooth muscle actin (αSMA), which is considered to be the best-known fibrosis marker. The aim of the study was to evaluate the expression of the αSMA in patients with PDAC and CP as the possible differentiation marker. Methods: We enrolled 114 patients undergoing pancreatic resection: 83 with PDAC and 31 with CP. Normal fragments of resected specimen from 21 patients represented the control tissue. The immunoexpressions of αSMA were detected in tissue specimens with immunohistochemistry (Abcam antibodies, GB). Results: Mean cytoplasmatic expression of αSMA protein in PDAC stromal cells was significantly higher compared to CP: 2.42 ± 0.37 vs 1.95 ± 0.45 (p < 0.01) and control group 0.61 ± 0.45 (p < 0.01). Strong immunoexpression of the αSMA protein was found in the vast majority (80.7%) of patients with PDAC, in about half (58%) of patients with CP, and not at all in healthy tissue. The expression of αSMA of different intensity was found in all patients with PDAC and CP, while in healthy tissue was minimal or absent. In PDAC patients, αSMA expression was significantly higher in tumors of diameter higher than 3 cm compared to smaller ones (p = 0.017). Conclusions: Presented findings confirm the significant role of fibrosis in both PDAC and CP; however, they do not confirm the role of αSMA as a marker of differentiation.

Attenuation of Expression of Splenocytes Pro-Inflammatory Cytokines and Lung Alpha-Smooth Muscle Actin By Human Amniotic Mesenchymal Stem Cells-Conditioned Medium in Asthmatic Mice

Background Asthma is a chronic respiratory illness characterized by lung tissue remodeling, T helper cell imbalance, and the generation of inflammatory factors. The Human amniotic mesenchymal stem cells-conditioned medium (hAM-MSC-CM) contains various immunomodulatory components and has been utilized in certain studies as a source for anti-inflammatory factors. We investigated the impacts of CM on splenocytes pro-inflammatory cytokines and alpha-smooth muscle actin (α-SMA) expression in Balb/c mice with Ovalbumin (OVA)-induced asthma.Methods and results Forty mice were separated into four groups of ten: control (challenged and sensitized with normal saline solution), asthma (sensitized on days 1, 8, and 14 and challenged daily with OVA from days 21 to 28), OVA+CM (asthmatic mice treated with CM on days 29 and 30), and OVA+DMEM (DMEM-treated asthmatic mice on days 29 and 30). The spleen and lung tissues were removed 48 hours after the final challenge, and the expression of the inflammatory factors in the splenocyte culture supernatant was determined by ELISA, while the α-SMA expression in the lung was determined using western blotting. α-SMA protein expression was significantly greater in lung tissue. Also, inflammatory agents were significantly higher in the splenocytes supernatant in the DMEM and asthma groups compared to the control group. CM therapy has been shown to inhibit the production of the α-SMA protein and inflammatory cytokines. Conclusions Results showed that CM Treatment was able to decrease the α-SMA expression in lung and splenocytes pro-inflammatory cytokines.