Long Non-Coding RNA TUG1 Gene Polymorphism and TUG1 Expression Level as Molecular Biomarkers of Systemic Lupus Erythematosus and Lupus Nephritis.

Long non-coding RNA (lncRNA) TUG1 acts as a proto-oncogene, allowing the proliferation of tumor cells, and it has been related to inflammation. Therefore, we aimed in this study to investigate for the first time the role of TUG1 gene polymorphism and the TUG1 level as biomarkers in systemic lupus erythematosus (SLE) and their link to lupus nephritis 145 SLE. A total of 145 healthy controls were subjected to clinical and laboratory evaluation. The disease activity was assessed by the SLE disease activity index (SLEDAI) score. SLE patients were divided into two subgroups according to the presence of lupus nephritis. The TUG1 gene polymorphisms rs5749201 and rs886471 were determined by Sanger sequencing, and TUG1 expression was assessed by qRT-PCR. There was a significant increase in the risk of SLE AA, TA, dominant genotypes, and the A allele of rs5749201 (p < 0.001) by 4.9-, 10.1-, 6.5-, and 2.5-fold in comparison to the relative control. GG and TG, dominant genotypes and the G allele of rs886471 (p < 0.01) increased the risk by 5.09-, 11.9-, 6.5-, and 2.6-fold. AA, A allele, dominant and recessive rs5749201genotypes increased the risk of lupus nephritis by 16.6-, 7.4-, 7.1-, and 12.2-fold, respectively (p < 0.05). GG, dominant and recessive genotypes, and the G allele of rs886471 increased the risk of lupus nephritis by 17.04-, 7.8-, 9.4-, and 6.08-fold, respectively (p < 0.05). Additionally, the AG haplotype increased the risk of SLE and lupus nephritis by 2.7- and 7.8-fold, respectively. The AA rs5749201 and GG rs886471 variants are significantly associated with more severe disease (p < 0.001). TUG1 expression was significantly higher in SLE than in the control and in the lupus nephritis than in non-lupus nephritis cases (p < 0.05). Interestingly, AA rs5749201 and GG rs886471 were significantly associated with higher TUG1 levels (p < 0.001). It was also found that AA rs5749201 and high SLEDAI were predictors of lupus nephritis. Overall, our findings illustrated for the first time that TUG1 gene rs5749201 and rs886471 variants were associated with increased risk of SLE, more severe disease, and lupus nephritis, and the TUG1 level could be used as a diagnostic biomarker of SLE and lupus nephritis.

Curcumin Nanofiber PCL/PLGA/Collagen Enhanced the Therapeutic Efficacy of Mesenchymal Stem Cells against Liver Fibrosis in Animal Model and Prevented its Recurrence.

The aim of this study is preconditioning of hBM-MSCs using curcumin modified nanomembrane to optimize therapy of hepatic fibrosis and preventing its recurrence. Methods: The nanomembrane was prepared by electrospinning technique and characterized using conventional method (cur- nanoscaffold and cur+ nanoscaffold). Kinetic release of curcumin was also measured by spectrophotometry. MSCs were isolated from human bone marrow (hBM-MSCs) and cultured on the both nanoscaffolds. We evaluated the in-vivo effect of hBM-MSCs from both nanoscaffold cultures (cur- nanoscaffold/hMSCs and cur+ nanoscaffold/MSCs) on liver fibrosis from its effective and preventive points and we assessed the mechanisms of these effects as in vitro studies as cell proliferation, its effect on hepatogenic differentiation, its effect on paracrine release of hBM-MSCs and in-vivo studying the effect on cell migration, survival, engraftment, fate of transplanted cells, modifying the fibrogenic and inflammatory microenvironments. Results: The results of animal model showed that single injection of preconditioning of hBM-MSCs using curcumin modified nanoscaffold ameliorate the fibrosis and prevent its recurrence until 24 weeks of therapy in contrast to improvement but not ameliorative effect of hBM-MSCs/ curcumin negative nanoscaffold which recurred progressively after 12 weeks of therapy. These effects of curcumin modified nanoscaffold were results from its highly efficacy on cell proliferation, in-vitro and in-vivo hepatogenic differentiation, increasing cell migration, engraftment and survival in the inflammatory microenvironment which was markedly improved by down regulation of inflammatory mediators and upregulation of anti-oxidant factors. Conclusion: hBM-MSCs cultured on the prepared curcumin nanomembrane in this study is promising in regenerative therapy for ameliorating the hepatic fibrosis and to prevent its recurrence.

Curcumin preconditioned mesenchymal stem cells derived exosomes transplantation ameliorate and protect against non- alcoholic steatohepatitis by regulation the expression of key genes of inflammation and oxidative stress.

BACKGROUND: Mesenchymal stem cells (MSCs) derived exosomes (MSCs/Exo) is considered a new strategy in cell free regenerative therapy. Curcumin preconditioning of MSCs reported to improve the anti- inflammatory and immunomodulatory properties of MSCs. We investigated the efficacy of exosome (Exo) obtained from curcumin-preconditioned MSCs (MSCs/Exo-Cur) vs. MSC/Exo without curcumin to ameliorate and prevent recurrence of non-alcoholic fatty liver (NASH) disease. METHODS AND RESULTS: In-vivo, methionine/choline-deficient diet (MCD) induced mice non-alcoholic fatty liver disease (NASH) were injected with MSCs/Exo without curcumin or MSCs/Exo-Cur with curcumin. We found that mice treated with MSCs/Exo-Cur had significantly ameliorated steatosis, inflammation, as evaluated by the reduced fibrosis in histopathological examination, decreased the serum level of liver enzymes (p < 0.001), liver triglycerides (TG) (p < 0.001) and cholesterol (Ch) (p < 0.001) and increased the lipid peroxidation (p < 0.001) compared to MSCs/Exo-treated mice. These effects remained for 3 months after treatment in MSCs/Exo-Cur-treated mice while features of NASH returned in MSCs/Exo-treated group. In vitro, HepG2 cells were cultured with palmitic acid (PA) and treated with MSCs/Exo or MSCs/Exo-Cur: the MSCs/Exo-Cur exposure reversed the lipotoxic effect from 4.5 to 1.7 fold vs 4.0 fold in MSCs/Exo and oxidative stress in PA-treated HepG2 cells (p < 0.001). We found that MSCs/Exo-Cur regulated the key markers of inflammatory and oxidative stress, genes responsible for fibrogenesis of the liver, key genes of lipid synthesis and transport. Interestingly, MSCs/Exo-Cur significantly down regulated the ASK-JNK-BAX genes involved in mitochondrial stress and apoptosis compared to MSCs/Exo (p < 0.001). CONCLUSION: Our study indicated that exosomes derived from curcumin preconditioned MSCs were able to ameliorate and protect against recurrence of NASH and regulated inflammatory, oxidative stress and mitochondrial-dependent apoptosis ASK-JNK-BAX genes.

A Novel Construct of Coral Granules-Poly-L-Lactic Acid Nanomembrane Sandwich Double Stem Cell Sheet Transplantation as Regenerative Therapy of Bone Defect Model.

OBJECTIVES: We examined the use of a new approach in nanotechnology and stem cell research as regenerative therapy for bone tissue defects. MATERIALS AND METHODS: We compared in vitro osteogenic potential of human Wharton jelly mesenchymal stem cells using coral granules and poly-L-lactic acid nanofiber according to proliferation (by cck-8 kit) and osteogenes (runt-related transcription factor 2, alkaline phosphatase, osteonectin) by quantitative reverse transcription-polymerase chain reaction, alkaline phosphatase assay, calcium measurement, and assessment of mineralization by Alizarin red and von Kossa staining. To overcome the limitations of natural coral, we made a modification by packaging the coral granules-human Wharton jelly mesenchymal stem cells by nanomembrane-human Wharton jelly mesenchymal stem cells to form sandwich double cell sheets and compared this hole with other holes (one was filled by human Wharton jelly mesenchymal stem cell suspension, and the other was filled by coral granules saturated with preinduced mesenchymal stem cells) by radiological and histopathological studies for repairing the bone gap. RESULTS: Collagen-coated poly-L-lactic acid showed higher mRNA levels for all osteogenes (P < .001), higher alkaline phosphatase and calcium content (P < .001), and greater stainability. Our in vivo experiment showed that the holes implanted with sandwich double cell sheet-poly-L-lactic acid coral were completely filled mature compact bone. The holes implanted with human Wharton jelly mesenchymal stem cells alone were filled with immature compact bone. Holes implanted with coral granules-human Wharton jelly mesenchymal stem cells were filled with condensed connective tissue. CONCLUSIONS: Poly-L-lactic acid nanofiber has greater osteogenic differentiating effect than the coral granules. The new approach of sDCS-PLLA-coral construct proved success for bone regeneration and repairing the bone gap and this may improve the design of tissue constructs for bone tissue regenerative therapy.

A novel function of collagen/PCL nanofiber interaction with MSCs in osteoarthritis is potentiation its immunomodulatory effect through increased ICAM expression.

BACKGROUND: The previous studies investigated the scaffold as mechanical carrier to stem cells or growth factors as transplantation therapy. This study aimed to investigate the effect of nanofiber scaffold on immunomodulatory properties of MSCs in osteoarthritis (OA) and the mechanism of this effect. METHODS: Collagen coated PCL nanofiber was prepared and examined by SEM. Mononuclear cells (MNCs) from the peripheral blood (P.B) and synovial fluid (SF) cultured with BMMSCs after stimulation by PHA using two types of co-culture (MSCs/Nano- and MSCs/nano+). Using flowcytometry, CD4, CD8, CD28, activation markers CD38 and CD23, regulatory CD4CD25high foxp3, CD14, IL17 by ELISA and cell proliferation using CCK-8 were assessed. ICAM mRNA was assessed by qRT-PCR and flowcytometry. RESULTS: MSCs/nano+ showed more significant inhibitory effect of MSCs regarding CD4, CD8, CD28, CD28 MFI, CD38, CD38 MFI, CD23, CD23 MFI, CD14, IL17 (p < 0.001) and higher level of Treg cells (p < 0.001) than nano- with difference regarding cell proliferation (p = 0.275). Also, the immunomodulatory effect of MSCs culture on nano+ on lymphocyte separated from S·F of osteoarthritis patients more powerful than nano-. The secretory function of MSCs culture on nano+ is more beneficial than that of nano-. Nano+ showed higher level of ICAM mRNA and CD54 than nano-. CONCLUSION: The present study proved a novel function of nanofiber by increasing the immunomodulatory effect of the MSCs in osteoarthritis through increased the ICAM expression and this promising for increase the success of MSCs transplantation in OA.

Association of pre-pro-endothelin gene polymorphism and serum endothelin-1 with intradialytic hypertension in an Egyptian population.

BACKGROUND: Intradialytic hypertension (IDH) is a major problem of hemodialysis and it is a multifactorial disorder and need early identification and management. AIM: Evaluate the serum concentration of endothelin-1 in patients with IDH and healthy control and the impact of pre-pro-endothelin gene polymorphism on level of endothelin-1 and susceptibility to IDH in Egyptian population. METHODS: The patient groups divided into group I, End stage renal disease (ESRD) on chronic hemodialysis with IDH (112); group II, ESRD on chronic hemodialysis without IDH (112); group III, healthy control (112). All undergone to full history, clinical examination, routine laboratory investigations, echocardiography, serum ET-1 level by ELISA and A(8002)G polymorphism detection in pre-pro-endothelin gene by PCR-RFLP. RESULTS: Our results showed significantly higher concentration of Endothelin-1 (ET-1) in both patient groups than healthy control and in group with IDH than cases without IDH (p < 0.001). GG, GA and mutated G allele carry the risk of IDH (OR = 15.94, 13.5, 5.51 respectively p < 0.001). There was association between GG and GA genotypes and higher ET-1 level in both patient groups (p < 0.001) and association between GG and GA genotype and higher mean arterial pressure (MAP), delta MAP (DMAP) and increased left ventricular mass index (LVMI) in both patient groups (p = 0.001, 0.028). CONCLUSION: Pre-pro-endothelin gene polymorphism A(8002)G is an independent risk factor for IDH through changing the level of ET-1 concentration in Egyptian population undergoing chronic hemodialysis.

HLA-G Gene Polymorphism in Egyptian Patients with Non-Hodgkin Lymphoma and its Clinical Outcome.

BACKGROUND: Non-Hodgkin lymphoma (NHL) is a major cancer in Egypt and worldwide and has many risk factors including genes involved in the immune response. AIM: we investigated the HLA-G 14bp gene polymorphism as a risk factor for NHL and its clinic pathologic features. The study involved 150 patients with NHL and 100 healthy control. Full histories, clinical examination, C.T scan and laboratory investigations such as CBC, LDH, ?2microglobulin and HCV RNA by qualitative real time PCR were performed for all subjects. HLA-G 14bp ins/del gene polymorphism was determined by PCR. RESULTS: in our study, del/del, ins/del and dominant genotypes increased the risk of NHL by 11.01, 10.55 and 10.88 fold respectively (p<0.001) but the recessive genotype did not increase the risk of NHL (p=0.112). Cases with the del allele had a greater risk of NHL than those with the ins allele (p<0.001). del/del and ins/del genotypes were significantly associated with higher LDH and ?2microglobulin levels (p<0.001), lower Hb and platelet values (p<0.001), extra nodal sites (p=0.001), poor performance status (p=0.04) and relapse (p=0.001).  Conclusions:  the results suggest that HLA-G 14bp ins/del gene polymorphism is a risk factor for NHL in our Egyptian population and is associated with poor clinical pathological features. ABBREVIATIONS: Non-Hodgkin lymphoma (NHL), Diffuse large B-cell lymphoma (DLBCL), follicular lymphoma (FL), Epstein-Barr virus (EBV), human T-cell lymphotropic/leukemia virus-1 (HTLV-1).

Modulation the expression of natural killer cell activating receptor (NKp44) in the peripheral blood of diffuse large B-cell lymphoma patients and the correlation with clinic pathological features.

NK cell activation is one strategy to improve the immunotherapy of non-Hodgkin's lymphoma. So, we aimed to investigate expression of Natural killer cell activating receptor NKp44 in patients with diffuse large B-cell lymphoma (DLBCL) and its correlation with clinic pathological data. In this study, 30 new cases with DLBCL in addition to 20 healthy control were involved. All were submitted to full history, clinical examination, histopathology, Routine laboratory investigations including CBC, LDH, ß2microgloubine and bone marrow examination. Cell culture of peripheral blood mononuclear cells and expression of CD56 and NKp44 by flowcytometry was done. We demonstrated increased NK cell populations (CD 56 +ve NKp44 -ve, CD 56 -veNKp44 +ve, total CD 56 +ve) and NKp44 MFI after in-vitro activation in both healthy control and DLBCL cases except for CD 56 +ve NKp44 +ve which significantly increased in patients not in healthy control (p=0.005, 0.601) respectively. No significant difference between the DLBCL and healthy control regarding all NK cell populations without PHA stimulation. However, the culture with PHA in DLBCL showed significant increase in NK cell populations than the healthy control (CD 56 +ve NKp44 +ve 12.37±7.52vs 6.80±4.07, p=0.008), (Total CD 56 +ve 18.80±8.74vs 12.66±5.17, p=0.017), (MFI of NKp44 10.95±6.18vs 5.58±1.70, p=0.001). Regarding the association with clinic pathologic features, increased expression of NKp44 was associated with lower values of LDH and earlier stages of DLBCL (p<0.05). So, activating receptor NKp44 can be modulated by in-vitro activation, hence improvement of its function as an approach of immunotherapy of DLBCL.