Potential probiotic Lactobacillus delbrueckii subsp. lactis KUMS-Y33 suppresses adipogenesis and promotes osteogenesis in human adipose-derived mesenchymal stem cell.

Today, probiotics are considered to be living microorganisms whose consumption has a certain number of beneficial effects on the consumer. The present study aimed to investigate the effect of a new probiotic extract (Lactobacillus delbrueckii subsp. lactis KUMS Y33) on the differentiation process of human adipose-derived stem cells (hADSCs) into adipocytes and osteocytes and, as a result, clarify its role in the prevention and treatment of bone age disease. Several bacteria were isolated from traditional yogurt. They were evaluated to characterize the probiotic's activity. Then, the isolated hADSCs were treated with the probiotic extract, and then osteogenesis and adipogenesis were induced. To evaluate the differentiation process, oil red O and alizarin red staining, a triglyceride content assay, an alkaline phosphatase (ALP) activity assay, as well as real-time PCR and western blot analysis of osteocyte- and adipocyte-specific genes, were performed. Ultimately, the new strain was sequenced and registered on NBCI. In the probiotic-treated group, the triglyceride content and the gene expression and protein levels of C/EBP-α and PPAR-γ2 (adipocyte-specific markers) were significantly decreased compared to the control group (P < 0.05), indicating an inhibited adipogenesis process. Furthermore, the probiotic extract caused a significant increase in the ALP activity, the expression levels of RUNX2 and osteocalcin, and the protein levels of collagen I and FGF-23 (osteocyte-specific markers) in comparison to the control group (P < 0.05), indicating an enhanced osteogenesis process. According to the results of the present study, the probiotic extract inhibits adipogenesis and significantly increases osteogenesis, suggesting a positive role in the prevention and treatment of osteoporosis and opening a new aspect for future in-vivo study.

Examining hepatoprotective effects of astaxanthin against methotrexate-induced hepatotoxicity in rats through modulation of Nrf2/HO-1 pathway genes.

Methotrexate (MTX), as a folic acid antagonist, is an effective drug in treating a wide range of malignancies and autoimmune diseases. However, the clinical use of MTX has been limited due to its side effects, the most common of which is hepatotoxicity. In this study, rats were randomly divided into six groups: three treatment groups received methotrexate and different doses of astaxanthin (AX) for 14 days. At the end of the study, blood samples were collected to determine serum levels of ALT, AST, ALP, and LDH. Also, liver tissues were isolated to evaluate antioxidant enzymes and markers of oxidative stress, histopathological damage, and expression of NF-E2-related transcription factor (Nrf2) and Heme oxygenase-1 (HO-1) genes. The results showed that administration of MTX significantly increased the levels of ALT, AST, ALP, and LDH in the blood, markers of oxidative stress, and histopathological damage in liver tissue and significantly reduced the levels of antioxidant enzymes and the expression of Nrf2 and HO-1 genes. On the other hand, treatment with AX decreased blood levels of ALT, AST, ALP, and LDH and oxidative stress markers and remarkably raises the activity of antioxidant enzymes and expression of Nrf2 and HO-1 genes in liver tissue. In addition, histopathological lesions were improved with AX administration. The findings of this study indicated that AX may be useful for the prevention of MTX-induced hepatotoxicity by improving oxidative and inflammatory changes.

Evaluation of the osteogenic effect of apigenin on human mesenchymal stem cells by inhibiting inflammation through modulation of NF-κB/IκBα.

Background and purpose: Apigenin has stimulatory effects on osteogenic differentiation of human mesenchymal stem cells (hMSCs) as well as anti-inflammatory properties. This study investigated the osteogenic differentiation of hMSCs in inflammatory conditions treated with apigenin focusing on nuclear factor kappa-light-chain-enhancer of activated B (NF-кB), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (IκBα) and nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain containing 3 (NLRP3) inflammatory pathways. Experimental approach: Along with osteogenic differentiation of the hMSCs, they became inflamed with lipopolysaccharide (LPS)/palmitic acid (PA) and treated with apigenin. Alizarin red staining, alkaline phosphatase (ALP) activity, and Runt-related transcription factor 2 (RUNX2) gene expression were used to determine the degree of differentiation. Also, gene expression of NLRP3 was performed along with protein expression of interleukin 1-beta (IL-1ß), NF-кB, and IκBα. Findings / Results: Apigenin was shown to be effective in neutralizing the inhibitory impact of LPS/PA on osteogenesis. Apigenin increased MSC osteogenic capacity by inhibiting NLRP3 expression and the activity of caspase-1. It was also associated with a considerable decrease in the protein expression of NF-κB and IκBα, as well as IL-1ß, in these cells. Conclusion and implications: The effects of apigenin on osteogenesis under inflammatory conditions were cautiously observed.

Intraprostatic injection of exosomes isolated from adipose-derived mesenchymal stem cells for the treatment of chronic non-bacterial prostatitis.

Mesenchymal stem cells (MSCs) own the capacity to secrete trophic factors as exosomes which play significant roles in regulating the functions of other cells and preventing inflammation. Due to the inflammatory process in chronic non-bacterial prostatitis (CNP) and the ambiguity in the treatment of this disease, the present study was aimed to investigate the therapeutic use of adipose-derived MSC exosomes in an animal model of CNP. MSCs were first isolated from rat subcutaneous adipose tissue, and exosomes were extracted from them. Specific features of exosomes were characterized by a scanning electron microscope, western blot technique, and Dynamic Light Scattering methods. To establish CNP in rats, intraprostatic injection of Freund's complete adjuvant was done. After confirmation of prostatitis, intraprostatic injections of exosomes were performed for treatment. Histological evaluation revealed that treatment with exosomes resulted in a relative improvement of lesions caused by CNP. The expression of p-NF-κB and p-IκBα proteins along with inflammatory markers was significantly increased in the CNP group, which treatment with exosomes significantly reduced their expression as well as IL-1ß and TNF-α proteins. The antioxidant effects of exosomes were also determined by significantly regulating glutathione peroxidase and superoxide dismutase activity and malondialdehyde levels in these animals. Our results cautiously suggest the therapeutic effects of MSC-derived exosomes against CNP-induced prostatitis through their antioxidant and anti-inflammatory activities, which should be further considered in the future.

Protective effects of combined Losartan and Nilotinib on carbon tetrachloride (CCl4)-induced liver fibrosis in rats.

Tyrosine kinase inhibitors (TKIs) have been developed as therapeutic compounds for inhibiting the progression of liver fibrosis. In the present study, the simultaneous treatment of Nilotinib (TKIs) and Losartan was studied. Forty rats were divided into eight groups of fibrosis induced by carbon tetrachloride (CCl4) and therapeutics (Nilotinib, Losartan, and combination therapy). In the end, serum parameters of the liver and gene expression analysis of transforming growth factor-ß1, its receptors (TßRII), platelet-derived growth factor, its receptors (PDGFRß), matrix metalloproteinases (MMP-2 and MMP-9), tumor necrosis factor-α, cytochrome P450 2E1, and collagen1 type 1 were performed. The oxidant/antioxidant factors were also analyzed. Histopathology analysis along with α-SMA immunohistochemistry and hydroxyproline evaluation was also conducted for a more in-depth study. The overall results indicated a better therapeutic effect of co-treatment of Nilotinib-Losartan in comparison with the treatment of each of them alone. Interestingly, some gene and protein factors and fibrotic indices were reduced even to the normal levels of the control group. The results of this study suggest that co-administration of these two combinations, strengthens their anti-fibrotic properties and, due to the routine use of these compounds against AML and blood pressure, these compounds can be used with caution against human liver fibrosis.

Hydrogen peroxide: a potent inducer of differentiation of human adipose-derived stem cells into chondrocytes.

Common protocols for chondrogenic differentiation of adipose-derived mesenchymal stem cells (ADSCs) are generally expensive and time-consuming and, so far, have not successfully recreated pure chondrocytes. We hypothesise that a low level of H2O2 may induce differentiation of ADSCs into chondrocytes in a shorter incubation time and relatively lower cost. Therefore, this study aimed to comparatively investigate the effectiveness of H2O2-containing or free medium in the induction of ADSCs to chondrocytes. ADSCs were isolated from the lipoaspirate of four healthy females and evaluated by immunophenotyping for their CD90, CD73, CD44, CD34, and CD45 cell surface markers. Chondrogenic differentiation was carried out using differentiation medium in the presence or absence of 10 and 50 µM H2O2 in normal and three-dimensional culture system. The intracellular contents of reactive oxygen species (ROS) were detected by flow cytometry and fluorescence microscopy. The hydroxyproline, was assessed as marker of collagen and the glycosaminoglycans (GAGs) content was both qualitatively detected and quantitatively determined. Real-time PCR was performed to determine the gene expression level of aggrecan (ACAN), type-II collagen, and transcription factor Sox9. H2O2-treated cells showed pre-chondrocyte morphology on day 1 and chondrocyte pellets were formed on day 14. H2O2-treated cells induced greater pellet sizes and showed significantly higher content of GAGs and hydroxyproline level compared with untreated cells. The gene expression levels of ACAN, collagen type-II, and Sox9 were markedly upregulated by H2O2. Our findings showed for the first time that H2O2-containing differentiation medium is potentially more effective than H2O2-free differentiation medium in the induction of chondrogensis of ADSCs.

The Role of Calcium in Differentiation of Human Adipose-Derived Stem Cells to Adipocytes.

Differentiation process of mesenchymal stem cells (MSCs) into adipocyte is involved in obesity. Multiple factors such as Ca2+ play important roles in different stages of this process. Because of the complicated roles of Ca2+ in adipogenesis, the aim of present investigation was to study the influx and efflux of Ca2+ into and out of the cells during adipogenesis. Adipose-derived MSCs were used to differentiate into adipocytes. MSCs were exposed to 2.5 mM Ca2+ or 1.8 mM Ca2+ plus calcium ionophore, A23187, for 3 days. Lipid staining, triglycerides (TG) content, and glyceraldehyde phosphate dehydrogenase (GAPDH) activity were evaluated to confirm the efficiency of the differentiation. Gene expression of GLUT4, PPARγ2, RAR-α, and calreticulin, as well as the protein levels of GLUT4 and PPARγ2 were determined. Ca2+ and in particular Ca2+ plus A23187 significantly lowered the efficiency of differentiation accompanied by decrease in intracellular TG deposits, GAPDH activity and alleviation of gene, and protein levels of GLUT4 and PPARγ2. While calreticulin and RAR-α were remarkably upregulated in A23187 group. This study showed the inhibitory effects of calcium in adipogenesis. Additionally, it indicated the greater inhibitory effect of calreticulin and RAR-α in controlling adipogenesis by higher levels of calcium.

Hepatoprotective effect of parthenolide in rat model of nonalcoholic fatty liver disease.

CONTEXT: The active ingredients of traditional medical herbs have been the focus of scientific interests. OBJECTIVE: This study was designed to explore the mechanisms of actions of parthenolide on nonalcoholic fatty liver disease (NAFLD). MATERIALS AND METHODS: Thirty-five male Wistar rats were fed high-fat diet (HFD) for eight weeks with or without an intraperitoneal injection of parthenolide to develop NAFLD. Liver triacylglycerol (TG), total antioxidant capacity (TAC), total oxidative status (TOS), thiobarbituric acid reactant substances (TBARs), total thiol groups and tumor necrosis factor alpha (TNF-α) and cytochrome P4502E1 (CYP2E1) levels as well as liver ALT, AST and catalase activities were determined. In addition, quantitative real-time PCR was performed to obtain hepatic gene expression levels of TNF-α, CYP2E1 and nuclear factor-κB (NF-κB). RESULTS: HFD caused a significant weight gain and increased liver TG content as well as alteration in ALT and AST activities, which were attenuated after administration of parthenoide (p < .05). Weakened liver antioxidant system (TAC, total thiol groups and catalase activity) and increased oxidative stress markers (TBARs and TOS) were mainly ameliorated by parthenolide treatment (p < .05). Increased hepatic TNF-α, NF-κB and CYP2E1 at the both gene expression and protein levels were found associated with necroinflammatory changes in histopathological observations and were abrogated almost completely after parthenolide treatment. Oxidative and inflammatory changes observed in HFD fed rats were indicative of NAFLD, which were suppressed with parthenolide treatment. CONCLUSIONS: Based on these results, parthenolide might be a candidate agent for preventing NAFLD due to its anti-inflammatory and anti-oxidative potency.

Dasatinib prevent hepatic fibrosis induced by carbon tetrachloride (CCl4) via anti-inflammatory and antioxidant mechanism.

OBJECTIVES: Dasatinib, a potent and broad-spectrum tyrosine kinase inhibitor, is approved for the treatment of imatinib-resistant chronic myelogenous leukemia. The aim of this study was to evaluate the anti-fibrotic, anti-inflammatory and antioxidant effects of this agent against CCl4-induced hepatic fibrosis and oxidative status. MATERIALS AND METHODS: Experimental fibrosis was induced in Wistar male rats by 12 weeks of CCl4 administration (i.p.). During the last 8 weeks of injection, rats were gavaged daily with Dasatinib (10 mg/kg). To evaluate anti-inflammatory and anti-fibrotic effects of Dasatinib, histopathological examination of liver tissue was performed and serum ALT and AST activities, oxidant, antioxidant parameters and hepatic tumor necrosis factor alpha (TNF-α) were examined. Moreover, transforming growth factor (TGF-ß1), platelet derived growth factor (PDGF) and TNF-α mRNA expressions were also evaluated by real time polymerase chain reaction. RESULTS: Dasatinib administration induced a significant reduction of ALT and AST activities (p < .001) and Malondialdehyde (MDA) content in CCl4 injected rats (p < .05). Concomitantly hepatic protein and mRNA expression of TNF-α, mRNA expression of TGF-ß1 and PDGF were increased due to CCl4 intoxication (p < .001), but Dasatinib treatment could significantly ameliorate these mediators at the level of gene expression (p < .01) and protein level of TNF-α (p < .001). The necro-inflammatory changes in histopathological finding, nitric oxide and hydroxyproline level were also increased during 12 weeks of CCl4 administration which was significantly attenuated by Dasatinib (p < .01). DISCUSSION AND CONCLUSION: Our findings indicate that Dasatinib can be cautiously an anti-fibrotic, anti-inflammatory and anti-oxidative agent in clinical setting.

Topical Application of Aloe vera Accelerated Wound Healing, Modeling, and Remodeling: An Experimental Study.

OBJECTIVE: Treatment of large wounds is technically demanding and several attempts have been taken to improve wound healing. Aloe vera has been shown to have some beneficial roles on wound healing but its mechanism on various stages of the healing process is not clear. This study was designed to investigate the effect of topical application of A. vera on cutaneous wound healing in rats. METHODS: A rectangular 2 × 2-cm cutaneous wound was created in the dorsum back of rats. The animals were randomly divided into 3 groups of control (n = 20), low-dose (n = 20), and high-dose (n = 20) A. vera. The control and treated animals were treated daily with topical application of saline, low-dose (25 mg/mL), and high-dose (50 mg/mL) A. vera gel, up to 10 days, respectively. The wound surface, wound contraction, and epithelialization were monitored. In each group, the animals were euthanized at 10 (n = 5), 20 (n = 5), and 30 (n = 10) days post injury (DPI). At 10, 20, and 30 DPI, the skin samples were used for histopathological and biochemical investigations; and at 30 DPI, the skin samples were also subjected for biomechanical studies. RESULTS: Aloe vera modulated the inflammation, increased wound contraction and epithelialization, decreased scar tissue size, and increased alignment and organization of the regenerated scar tissue. A dose-dependent increase in the tissue level of dry matter, collagen, and glycosaminoglycans' content was seen in the treated lesions, compared to the controls. The treated lesions also demonstrated greater maximum load, ultimate strength, and modulus of elasticity compared to the control ones (P < 0.05). CONCLUSIONS: Topical application of A. vera improved the biochemical, morphological, and biomechanical characteristics of the healing cutaneous wounds in rats. This treatment option may be valuable in clinical practice.