Repair of Osteochondral Defects in Rabbit Knee Using Menstrual Blood Stem Cells Encapsulated in Fibrin Glue: A Good Stem Cell Candidate for the Treatment of Osteochondral Defects

BACKGROUND: In recent years, researchers discovered that menstrual blood-derived stem cells (MenSCs) have the potential to differentiate into a wide range of tissues including the chondrogenic lineage. In this study, we aimed to investigate the effect of MenSCs encapsulated in fibrin glue (FG) on healing of osteochondral defect in rabbit model. METHODS: We examined the effectiveness of MenSCs encapsulated in FG in comparison with FG alone in the repair of osteochondral defect (OCD) lesions of rabbit knees after 12 and 24 weeks. RESULTS: Macroscopical evaluation revealed that the effectiveness of MenSCs incorporation with FG is much higher than FG alone in repair of OCD defects. Indeed, histopathological evaluation of FG + MenSCs group at 12 weeks post-transplantation demonstrated that defects were filled with hyaline cartilage-like tissue with proper integration, high content of glycosaminoglycan and the existence of collagen fibers especially collagen type II, as well as by passing time (24 weeks post-transplantation), the most regenerated tissue in FG + MenSCs group was similar to hyaline cartilage with relatively good infill and integration. As the same with the result of 12 weeks post-implantation, the total point of microscopical examination in FG + MenSCs group was higher than other experimental groups, however, no significant difference was detected between groups at 24 weeks (p>0.05). CONCLUSION: In summary, MenSCs as unique stem cell population, is suitable for in vivo repair of OCD defects and promising for the future clinical application.

Menstrual blood-derived stromal stem cells augment cd4+ t cells proliferation

Background: It is more than sixty years that the concept of the fetal allograft and immunological paradox of pregnancy was proposed and in this context, several regulatory networks and mechanisms have been introduced so far. It is now generally recognized that mesenchymal stem cells exert potent immunoregulatory activity. In this study, for the first time, the potential impact of Menstrual blood Stem Cells [MenSCs], as surrogate for endometrial stem cells, on proliferative capacity of CD4+ T cells was tested

Methods: MenSCs and Bone marrow Mesenchymal Stem Cells [BMSCs] were isolated and assessed for their immunophenotypic features and multi-lineage differentiation capability. BMSCs and MenSCs with or without IFNGamma pre-stimulation were co-cultured with purified anti-CD3/CD28-activated CD4+ T cells and the extent of T cell proliferation at different MenSCs: T cell ratios were investigated by CSFE flow cytometry. IDO activity of both cell types was measured after stimulation with IFNGamma by a colorimetric assay

Results: MenSCs exhibited dual mesenchymal and embryonic markers and multilineage differentiation capacity. MenSCs significantly increased proliferation of CD4+ cells at ratios 1:2, 1:4 and 1:8. IFNGamma pre-treated BMSCs but not MenSCs significantly suppressed CD4+ T cells proliferation. Such proliferation promoting capacity of MenSCs was not correlated with IDO activity as these cells showed the high IDO activity following IFNGamma treatment

Conclusion: Although augmentation of T cell proliferation by MenSCs can be a basis for maintenance of endometrial homeostasis to cope with ascending infections, this may not fulfill the requirement for immunological tolerance to a semi-allogeneic fetus. However, more investigation is needed to examine whether or not the immunomodulatory properties of these cells are affected by endometrial microenvironment during pregnancy

Current state of cartilage tissue engineering using nanofibrous scaffolds and stem cells

Cartilage is an avascular, aneural, and alymphatic connective tissue with a limited capacity caused by low mitotic activity of its resident cells, chondrocytes. Natural repair of full thickness cartilage defects usually leads to the formation of fibrocartilage with lower function and mechanical force compared with the original hyaline cartilage and further deterioration can occur. Tissue engineering and regenerative medicine is a promising strategy to repair bone and articular cartilage defects and rehabilitate joint functions by focusing on the optimal combination of cells, material scaffolds, and signaling molecules. The unique physical and topographical properties of nano?brous structures allow them to mimic the extracellular matrix of native cartilage, making an appropriate resemblance to induce cartilage tissue regeneration and reconstruction. To improve simulation of native cartilage, the incorporation of nanofibrous scaffolds with suitable corresponsive cells could be effective. In this review article, an attempt was made to present the current state of cartilage tissue engineering using nanofibrous scaffolds and stem cells as high proliferative immune privilege cells with chondrogenic differentiation ability. The comprehensive information was retrieved by search of relevant subject headings in Medline/Pubmed and Elsevier databases

Exogenous secreted frizzled-related protein-4 modulates steroidogenesis of rat granulosa cells through Wnt/beta-catenin and PI3K/AKT signaling pathways

Background: It has been reported that secreted frizzled-related protein-4 known as an antagonist of Wnt signaling pathway plays a role in luteinization process of rodent granulosa cells. The purpose of this study was twofold: 1] to determine whether recombinant human secreted frizzled-related protein-4 [rhSFRP-4] could directly induce terminal differentiation of rat Granulosa Cells [GCs] and 2] to understand how the modulation of beta-catenin and Protein Kinase B [PKB]/AKT activity by exogenous SFRP-4 could be involved in steroidogenesis

Methods: GCs were firstly stimulated with Follicle-Stimulating Hormone [FSH] named as FSH-primed cells then were treated with luteinizing hormone [LH]. Then estradiol [E[2]] and progesterone [P[4]] production levels were assessed in the absence or presence of rhSFRP-4 treatment. The expression levels of activated beta-catenin, pAKTser[473], pGSK3 beta ser[9] were assessed by western blot or immuno-fluoresence

Results: In the presence of rhSFRP-4, there was 38% decreased E[2] levels compared to untreated FSH-primed cells [p<0.05], and P[4] production subsequently decreased. However, in GCs pre-treated with rhSFRP-4 prior to addition of FSH, P[4] levels increased 2-fold compared with untreated cells [p<0.05]. Unexpectedly, treatment with rhSFRP-4 prior to LH stimulation inhibited LH-induced P[4] secretion. Treatment with low [0.5 ng/ml] but not high [50 ng/ml] concentrations of rhSFRP-4 led to significantly increased levels of pGSK3 beta ser[9] [1.6-fold] and nuclear active beta-catenin [2.8-fold] in GCs compared with untreated cells. Interestingly, pre-treating GCs with rhsFPR4 prior to LH stimulation resulted in a 38% decrease in pAKTser[473] levels compared with those in LH-treated cells [p<0.05]

Conclusion: Taken together, our results showed that rhSFRP-4 could directly induce terminal differentiation in GCs via the modulation of beta-catenin and PKB/AKT pathways and that it does so in a dose-dependent manner

Lipopolysaccharide- and lipoteichoic acid-mediated pro-inflammatory cytokine production and modulation of tlr2, tlr4 and myd88 expression in human endometrial cells

Toll-like receptor [TLR]-mediated inflammatory processes are supposed to be involved in pathophysiology of spontaneous abortion and preterm labor. Here, we investigated functional responses of human endometrial stromal cells [ESCs] and whole endometrial cells [WECs] to lipopolysaccharide [LPS] and lipoteichoic acid [LTA] Endometrial tissues were obtained from 15 cycling women who underwent laparoscopic tubal ligation. Modulation of TLR2, TLR4 and MyD88 expression and production of pro-inflammatory cytokines by WECs and ESCs in response to LPS and LTA were assessed. WECs and ESCs expressed significant levels of TLR4 and MyD88 transcripts but, unlike WECs, ESCs failed to express TLR2 gene. Regardless of positive results of Western blotting, ESCs did not express TLR4 at their surface as judged by flow cytometry. Immunofluorescent staining revealed intracellular localization of TLR4 with predominant perinuclear pattern. LPS stimulation marginally increased TLR4 gene expression in both cell types, whereas such treatment significantly upregulated MyD88 gene expression after 8 hr [p<0.05]. At the protein level, however, LPS activation significantly increased TLR4 expression by ESCs [p<0.05]. LTA stimulation of WECs was accompanied with non-significant increase of TLR2 and MyD88 transcripts. LPS and LTA stimulation of WECs caused significant production of IL-6 and IL-8 in a dose-dependent manner [p<0.05]. Similarly, ESCs produced significant amounts of IL-6, IL-8 and also TNF-alpha in response to LPS activation [p<0.05]. Our results provided further evidence of initiation of inflammatory processes following endometrial TLR activation by bacterial components which could potentially be harmful to developing fetus

Isolation and partial characterization of human amniotic epithelial cells: the effect of trypsin

Despite the extensive information available in the literature, cell surface marker signature of human Amniotic Epithelial Cells [hAECs] remains controversial. The aim of the present study was to characterize immunophenotypic features, proliferative capacity and immunogenicity of hAECs. We also tested whether expression of some cell surface markers is influenced by the type of trypsin used for tissue digestion. Single cell suspensions of amniotic membranes from four human placentas were isolated by enzymatic digestion and expression of CD9, CD10, CD29, CD34, CD38, CD44, CD45, CD73, CD105, CD133, HLA-I, HLA-DR, HLA-G, SSEA-4, STRO-1 and OCT-4 was then evaluated by flow cytometry. The differential impact of four trypsin types on the yield and expression of CD105 and HLA-I was also determined. The proliferative capacity of cultured hAECs was assessed and compared in the presence and absence of Epidermal Growth Factor [EGF]. To test their immunogenicity, hAECs were injected into Balb/c mice and the reactivity of hyperimmunized sera was examined by immunofluorescence staining. Nearly all purified cells expressed mesenchymal markers, CD9, CD10, CD29, and CD73 and the embryonic marker, SSEA-4. A large proportion of the cells also expressed STRO-1 and OCT-4. The purified cells also expressed HLA-G and HLA-I. A very small proportion of hAECs expressed CD34, CD38, CD44, CD133 and HLA-DR. The type of trypsin used for enzymatic digestion affected both the percentage and expression of HLA-I and CD105. hAECs revealed substantial proliferative capacity only when cultured in the medium supplemented with EGF. These cells were shown to be capable of inducing high amounts of anti-donor antibodies. Here we provided evidence that hAECs are immunogenic cells with high level of HLA-I expression. Furthermore, this work highlighted the impact of isolation procedure on the immunophenotype of hAEC

Expression of cytochrome P450 and glutathione S-transferase in human bone marrow mesenchymal stem cells

Currently several studies are being carried out on various properties of mesenchymal stem cells [MSCs] however there are a few investigations about drug metabolizing properties of these cells. The aim of this study was to measure the key factors involved in drug metabolism in human bone marrow MSCs. For this purpose, cellular glutathione [GSH], glutathione Stransferase [GSTs] and cytochrome P450 class 3A4 [CYP3A4] were detected in these cells. Results showed that CYP3A4 and GSTA1-1 mRNA are not detectable in MSCs however mRNA specific for GSTP1-1 was considerably expressed in MSCs. GSH content and GST activity was also detected in MSCs. These data suggest that human bone marrow MSCs possess the drug metabolizing activity which may be useful in handling drugs and chemotherapeutic agents passing to the bone marrow

Hepatic tissue engineering using scaffolds: state of the Art

Severe hepatic failure accounts for many deaths and raises medical costs each year worldwide. Currently, liver transplantation is the most common therapeutic option for patients with end-stage chronic liver disease. Due to decrease in the number of organ donors, many in need of transplantation continue to remain on the waiting list. Hepatic Tissue Engineering is a step toward alleviating the need for organ donors. Regenerative medicine and tissue engineering require two complementary bey ingredients as follows: 1] biologically compatible scaffolds that can be readily adopted by the body system without harm, and 2] suitable cells including various stem cells or primary cells that effectively replace the damaged tissues without adverse consequences. Yet many challenges must be overcome such as scaffold choice, cell source and immunological barriers. Today, hepatogenic differentiation of stem cells has created trust and promise for use of these cells in hepatic tissue engineering and liver replacement. However, using suitable scaffolds is an important bey to achieving the necessary functions required for hepatic replacement. In recent years, different scaffolds have been used for liver tissue engineering. In this review, we have presented different concepts in using cell /scaffold constructs to guide hepatic tissue engineering

Capillary network formation by endothelial cells differentiated from human bone marrow mesenchymal stem cells

Human bone marrow derived mesenchymal stem cells [HBMSCs] have the potential to differentiate into cells such as adipocyte, osteocyte, hepatocyte and endothelial cells. In this study, the differentiation of hBMSCs into endothelial like-cells was induced in presence of vascular endothelial growth factor [VEGF] and insulin-like growth factor [IGF-1]. The differentiated endothelial cells were examined for their ability to express VEGF receptor-2 [VEGFR2] and von willebrand factor [vWF]. Then the cells were adopted to grow and develop capillary network in a semisolid gel matrix in vitro. The capillary network formation in a well of 24-well plate was found to be 85% in presence of VEGF [50ng/ml] and IGF-1 [20ng/ml] of the culture media. These data may suggest that the expression of endothelial markers in endothelial like-cells derived from hBMSCs is associated with their ability to form capillaries

Development of a novel three-dimensional biocompatible nanofibrous scaffold for the expansion and hepatogenic differentiation of human bone marrow mesenchymal stem cells

In this present study, we examined the differentiation potential of human bone marrow derived mesenchymal stem cells [hBMSCs] into hepatocytes on a three-dimentional [3D] nanofibrous scaffold formed by Poly [e-caprolactone] [PCL], collagen and polyethersulfone [PES]. The nanofiber was prepared by the electrospining technique. HBMSCs were isolated using combining gradient density centrifugation with plastic adherence. Flow cytometric analysis was used to identify the isolated MSCs. The performance of the cells on the scaffold was evaluated by scanning electron microscopy [SEM] and MTT assay. The hBMSCs were then cultured in a hepatic differentiation medium containing hepatocyte growth factor [HGF], oncostatin M [OSM] and dexamethasone [DEX] for up to 21 days. The results showed that the isolated hBMSCs expressed specific markers such as CD44, CD166, CD105 and CD13. The integrity of the MSCs was further confirmed by their differentiation potential to osteogenic and adipogenic lineages. Scanning electron micrographs and MTT analysis revealed that the cells adhered and proliferated well on the nanofibrous hybrid scaffolds. Immunocytochemical analysis of albumin and a-fetoprotein [AFP] showed the accumulation of these markers in the differentiated cells on the scaffold. Hepatocyte differentiation was further confirmed by showing expression of albumin, AFP and cytokeratin-19 [CK-19] at mRNA levels in differentiated cells. In conclusion, the evidences presented in this study show that the engineered scaffold is promising for maintenance of hepatocyte-like cells suitable for transplantation

Class-Pi of glutathione S-transferases

Class-Pi of glutathione s-transferases [GST-Pi] is the specific form of GSTs that are known to participate particularly in the mechanisms of resistance to drugs and carcinogens. This class of the enzyme is referred to as class-P or class-Pi or class pi. The accepted terminology in this review article is class-Pi. In this article following a brief description of identified molecular forms of GSTs, we focus on GST-Pi. We review new findings about the structure and regulation of GST-Pi gene. Then, the role of GST-Pi in liver damage, oxidative stress, carcinogenesis and drug resistance are discussed. Also, the presence of common genetic polymorphism, hypermethylation in GST-Pi gene and the consequences GST-Pi knock out is regarded