Differentiation of Human Wharton Jelly Mesenchymal Stem Cells into Germ-Like Cells; emphasis on evaluation of Germ-long non-coding RNAs.

BACKGROUND: The proliferation and differentiation of stem cells into Germ-Like Cells (GLCs) is mediated by several growth factors and specific genes, of which some are related to long non-coding RNAs (lncRNAs). We have developed a modified differentiation process and identified a panel of GermlncRNAs related to GLCs. METHODS: Human Wharton Jelly Mesenchymal Stem Cells were treated with 25 ng/ml Bone Morphogenetic Protein (BMP)-4 and 10- 5 M all-trans retinoic acid to differentiate them into germ-like cells. To confirm the differentiation, changes in the expression of Oct-4, C-kit, Stella, and Vasa genes were assessed using quantitative Real-Time PCR (qPCR) and immunocytochemistry. QPCR was also used before and after differentiation to evaluate the changes in a lncRNA panel, using a 96-well array. Statistical analysis of the data was performed by SPSS 21. RESULTS: After 21 days of induction, the HWJ-MSCs derived germ-like cells were formed. Also, qPCR and immunocytochemistry showed that the pluripotent Oct4 marker was expressed in the undifferentiated HWJ-MSCs, but its expression gradually decreased in the differentiated cells. C-kit was expressed on days 7, 14, and 21 of differentiation. Both GLC markers of Stella and Vasa genes/proteins were present only in differentiated cells. Of the 44 lncRNA genes array, 36 of them showed an increase and eight genes showed a decrease. CONCLUSION: Our study showed that BMP4 and RA are effective in inducing HWJ-MSCs differentiation into GLCs. In addition, our study for the first time showed changes in the lncRNAs expression during the differentiation of HWJ-MSCs into GLCs by using BMP4 and RA.

Serum level and tumor tissue expression of Ribonucleotide-diphosphate Reductase subunit M2 B: a potential biomarker for colorectal cancer.

BACKGROUND: This study explored the applicability of serum level and tissue expression of Ribonucleotide-diphosphate Reductase subunit M2 B (RRM2B) as reliable biomarkers for colorectal cancer (CRC) progression and metastasis. METHODS AND RESULTS: The present descriptive-analytic cohort study was conducted on 50 newly diagnosed CRC patients (stage II, III) and 50 healthy individuals. The new cases had not received any therapeutic intervention and underwent surgery immediately after the initial diagnosis. Tumorous tissues and marginal healthy tissues (as control) were excised to determine the mRNA tissue expression of RRM2B by Real-Time PCR. Serum RRM2B protein was measured using an ELISA method once in the control group. In the patients, serum RRM2B protein was evaluated before, 1 and 3 months after surgery. The tumor metastasis node (TMN) classification system and liver metastasis were evaluated in CRC patients. The results showed significantly lower RRM2B serum levels in 1 and 3 months after surgery compared with the pre-surgery condition (P = 0.014, P < 0.001 respectively). The mean RRM2B gene expression was 51% lower in tumor tissue than its adjacent normal tissue (P < 0.001). No significant relationship was found between serum level of RRM2B and tumor staging and metastasis in patients before surgery (P = 0.373, P = 0.189), 1 month after surgery (P = 0.960, P = 0.088), and 3 months after surgery (P = 0.407, P = 0.724). RRM2B expression in tumor tissue is not associated with tumor staging and metastasis (P = 0.254, P = 0.721). CONCLUSION: These data suggest measuring serum protein level of RRM2B could have a role in CRC progression, although this study should be considered preliminary due to small sample size and short follow-up duration.

The lower expression of circulating miR-210 and elevated serum levels of HIF-1α in ischemic stroke; Possible markers for diagnosis and disease prediction.

BACKGROUND: Stroke, either due to ischemia or hemorrhage, causes acute neurological damages to the brain. There is shortage of reliable biomarkers for ischemic stroke (IS), and we therefore investigated the serum concentrations of microRNA-210 (miR-210) and hypoxia inducible factor-1α (HIF-1α), as possible diagnostic and/or prognostic markers for IS. METHODS: Serum samples were acquired from 52 IS patients and their healthy counterparts at five time points: upon admission, 24 and 48 h after admission, upon discharge and 3 months later. Serum levels of miR-210 and HIF-1α were respectively analyzed using real time RT-PCR and ELISA. Diagnostic and prognostic accuracy tests were performed to assess the value of suggested biomarkers. RESULTS: IS patients demonstrated higher levels of serum HIF-1α and lower miR-210 in comparison to the healthy subjects. MiR-210 was suggested to be a weak diagnostic biomarker at the time of admission (AUC = 0.61; p = 0.05), while HIF-1α was an acceptable diagnostic marker for IS (AUC = 0.73; p < 0.0001). The higher expression of miR-210 and lower levels of HIF-1α were associated with better survivals in IS patients. CONCLUSIONS: Serum miR-210 is a weak diagnostic marker of IS. Serum HIF-1α is a better biomarker in diagnosing IS patients but further work in larger groups, including those with hemorrhagic stroke is necessary to confirm its diagnostic utility. Similarly, the prognostic potentiality of miR-210 and HIF-1α was acceptable but needs bigger sample size and longer follow-up to be statistically confirmed.

The elevation of S100B and downregulation of circulating miR-602 in the sera of ischemic stroke (IS) patients: the emergence of novel diagnostic and prognostic markers.

Ischemic stroke (IS) is a major cause of mortality and disability. However, no reliable prognostic or diagnostic biomarker has been utilized to date. Here, we have evaluated the serum S100B concentration and miR-602 expression as potential biomarkers for IS. Fifty-two IS patients and 52 age- and sex-matched healthy volunteers were enrolled. Blood samples were collected from all patients at the time of admission, 24 and 48 h later, at the time of discharge, and 3 months later. Real-time (RT) PCR was used to measure the serum level of miR602. We also measured the serum concentration of S100B using ELISA. As compared with healthy subjects, IS patients had a higher level of serum S100B and lower serum miR-602. ROC curve analyses revealed that miR-602 (AUC = 0.8168; P < 0.0001) and S100B (AUC = 0.8699; P < 0.0001) had acceptable ability to differentiate between IS patients from healthy subjects. Furthermore, serum S100B was a reliable predictor of the survival outcome at 3 months (P = 0.021). The expression of miR-602 was significantly higher in patients with bigger NIHSS scores. The lower levels of miR-602 and higher concentration of S100B in the sera of IS patients could be associated with clinically significant diagnostic utilities. S100B could be also introduced as a reliable prognostic marker for stroke and implemented in future research.

Overexpression of microRNA-375 and microRNA-122 promotes the differentiation of human induced pluripotent stem cells into hepatocyte-like cells.

MicroRNAs (miRNAs) are involved in the regulation of gene expression. In this study, we evaluated the use of overexpression of microRNA-375 (miR-375) and miR-122 in differentiating the Human Induced Pluripotent Stem Cells (hiPSCs) into functional hepatocyte-like cells (HLCs) without growth factors. We also compared the differentiation by miRNAs versus growth factors. HiPSCs were divided into two main groups: 1- HiPSCs were induced using lentiviral overexpression of miR-375 to differentiate into definitive endoderm (DE) cells in seven days. Then lentiviral overexpression of miR-122 was applied to differentiate DE cells into HLCs in additional 14 days. 2- HiPSCs were differentiated into HLCs using growth factors in 21 days. DE and hepatocyte markers were investigated by qRT-PCR, immunofluorescence, secretion analysis and LDL uptake assay. In the produced cells of both groups: the expression levels of DE markers (FOXA2 and SOX17) and hepatocyte markers (albumin, CK18, and HNF4a) in comparison with the undifferentiated hiPSCs increased significantly in seven and 21 days respectively. The albumin and urea secretion and LDL uptake were also detected. These results weren't significantly different between two groups. Therefore, we demonstrated that the over expression of miR-375 and then miR-122 could differentiate hiPSCs into functional HLCs without growth factors for developing cell-based therapies.

Hybrid poly-l-lactic acid/poly(ε-caprolactone) nanofibrous scaffold can improve biochemical and molecular markers of human induced pluripotent stem cell-derived hepatocyte-like cells.

A suitable alternative strategy for liver transplantation is the use of nanofibrous scaffolds together with stem cells. In this study, a random hybrid of poly-l-lactic acid (PLLA) and poly(ε-caprolactone) (PCL) was used as a three-dimensional (3D) culture for differentiation of hepatocyte-like cells and compared with routine culture (two-dimensional [2D]). The expression of the endodermal marker, forkhead box A2 (FOXA2), was assessed on Day 3 and the hepatic markers; albumin (ALB), α-1 antitrypsin (AAT), and cytokeratin-18 (CK-18) were evaluated on Day 18 using quantitative polymerase chain reaction qPCR. As well as, ALB, α-fetoprotein (AFP), and low-density lipoprotein (LDL) uptake were evaluated using immunocytochemistry; moreover, periodic acid-Schiff and Oil Red were done by cell staining. In addition, AFP and urea production were evaluated by chemiluminescence and colorimetric assays. Light and scanning electron microscopy (SEM) showed changes in the cells in 2D and 3D models. The gene expression of hepatic markers was significantly higher in the 3D cultures. In addition, immunocytochemistry and cell staining showed that ALB, AFP, LDL-uptake, periodic acid-Schiff, and Oil Red were expressed in both cells derived on 2D and 3D. Furthermore, the evaluation of AFP and urea secretion was significantly different between 2D and 3D strategies. These findings suggest that functionally cells cultured on a PLLA/PCL scaffold may be suitable for cell therapy and regenerative medicine.

The Association between Inflammatory Markers in the Acute Phase of Stroke and Long-Term Stroke Outcomes: Evidence from a Population-Based Study of Stroke.

BACKGROUND: Little is known about the association between inflammatory markers in the acute stroke phase and long-term stroke outcomes. METHODS: In a population-based study of stroke with 5 years follow-up, we measured the level of serum heat shock protein 27 immunoglobulin G antibody (anti-HSP27), C-reactive protein (CRP), and pro-oxidant antioxidant balance (PAB) in the acute stroke phase. We analyzed the association between these inflammatory biomarkers and stroke outcomes (recurrence, death and disability/functional dependency) with using multivariable Cox proportional hazard models. RESULTS: Two hundred sixty-five patients with first-ever stroke were included in this study. The severity of stroke at admission, measured by National Institute of Health Score Scale was associated with serum concentration of CRP (Spearman's rank correlation coefficient rs = 0.2; p = 0.004). CRP also was associated with 1-year combined death and recurrence rate ([adjusted hazard ratio 1.06, 95% CI 1.01-1.12; p = 0.02]). However, we did not find any association between the concentrations of CRP, anti-HSP27, PAB, and 5-year death and stroke recurrence rates. None of 3 biomarkers was associated with the long-term disability rate (defined as modified Rankin Scale >2) and functional dependency (defined as Barthel Index <60). CONCLUSION: CRP has a significant direct, yet weak, correlation to the severity of stroke. In addition, the level of CRP at admission may have a clinical implication to identify those at a higher risk of death or recurrence.

MicroRNA-129-1 acts as tumour suppressor and induces cell cycle arrest of GBM cancer cells through targeting IGF2BP3 and MAPK1.

BACKGROUND: MicroRNA-129-1 (miR-129-1) seems to behave as a tumour suppressor since its decreased expression is associated with different tumours such as glioblastoma multiforme (GBM). GBM is the most common form of brain tumours originating from glial cells. The impact of miR-129-1 downregulation on GBM pathogenesis has yet to be elucidated. METHODS: MiR-129-1 was overexpressed in GBM cells, and its effect on proliferation was investigated by cell cycle assay. MiR-129-1 predicted targets (CDK6, IGF1, HDAC2, IGF2BP3 and MAPK1) were also evaluated by western blot and luciferase assay. RESULTS: Restoration of miR-129-1 reduced cell proliferation and induced G1 accumulation, significantly. Several functional assays confirmed IGF2BP3, MAPK1 and CDK6 as targets of miR-129-1. Despite the fact that IGF1 expression can be suppressed by miR-129-1, through 3'-untranslated region complementary sequence, we could not find any association between IGF1 expression and GBM. MiR-129-1 expression inversely correlates with CDK6, IGF2BP3 and MAPK1 in primary clinical samples. CONCLUSION: This is the first study to propose miR129-1 as a negative regulator of IGF2BP3 and MAPK1 and also a cell cycle arrest inducer in GBM cells. Our data suggests miR-129-1 as a potential tumour suppressor and presents a rationale for the use of miR-129-1 as a novel strategy to improve treatment response in GBM.

MiRNA-375 promotes beta pancreatic differentiation in human induced pluripotent stem (hiPS) cells.

Islet transplantation is considered as an ultimate option for the treatment of type I diabetes. Human induced pluripotent stem cells (hiPSCs) have raised the possibility that patient-specific insulin-secreting cells might be derived from somatic cells through cell fate reprogramming. However, current protocols mostly rely on the use of several cytokines and inhibitors for directing differentiation towards pancreatic fate. Given the high manufacturing cost of these recombinant proteins, this approach is prohibitive for clinical applications. Knowing that microRNAs (miRNAs) are key players in various stages of pancreatic development, we present a novel and cost-effective strategy in which over-expression of miR-375 promotes pancreatic differentiation in hiPSCs in the absence of any other stimulator. We used a polycistronic viral vector expressing Sox2, Klf4, c-Myc, and Oct4 to drive hiPSCs from human foreskin fibroblasts. The established hiPSCs are similar to human embryonic stem cells in many aspects including morphology, passaging, surface and pluripotency markers, and gene expression. For differentiation induction, miR-375 was lentivirally overexpressed in these hiPSCs. Morphological assessment, immunocytochemistry, and expression analysis of islet marker genes confirmed that islet like cells were obtained in miR-375 transduced cells compared to controls. Our differentiated clusters secreted insulin in a glucose-dependant manner, showing in vitro functionality. We demonstrated for the first time that miRNAs might be ideal substitutes to induce pancreatic differentiation in hiPSCs. This work provides a new approach to study the role of miRNAs in pancreatic specification and increase the feasibility of using patient-specific iPSCs for beta cell replacement therapy for type I diabetes.

Prooxidant-antioxidant balance and hs-CRP in patients with beta-thalassemia major.

BACKGROUND: Beta-thalassemia major is one of the most frequent hemoglobinopathies in the world. This is a study on the levels of prooxidant-antioxidant balance (PAB) that can measure the pro-oxidant burden and the anti-oxidant capacity simultaneously in one assay, giving a redox index, as well as an assessment of high-sensitivity C-reactive protein (hs-CRP) in patients with beta-thalassemia major. It also determines the impact both on kidney function tests. METHODS: Ninety subjects with beta-thalassemia major (betaTM) and 90 more healthy subjects as the control group were enrolled in the study. Laboratory parameters, serum hs-CRP, and the novel assay of prooxidant-antioxidant balance were measured. RESULTS: Biochemical and hematological variables, except MCHC, were significantly decreased in patients. Ferritin (p < 0.001) and PAB (p < 0.001) values were statistically increased in patients, whereas hs-CRP (p > 0.05) did not significantly increase in patients. Spearman's correlation univariate analysis showed no significant correlation between PAB with hs-CRP or ferritin. However, there was a significant statistical correlation between hs-CRP with creatinine and triglycerides. Also we found a slight and negative correlation between high-density lipoprotein cholesterol, hemoglobin, and hemotocrit with inflammatory factor. CONCLUSIONS: Briefly, we found higher levels of PAB values and a slight increase in hs-CRP concentration in patients, which may be involved in the pathological consequences of beta-thalassemia major and contribute to the gradual development of injury to the organs.

Association between Pro-oxidant-Antioxidant balance and high-sensitivity C-reactive protein in type 2 diabetes mellitus: A Study on Postmenopausal Women.

INTRODUCTION: Oxidative stress known as a predictive marker for cardiovascular and metabolic diseases could be measured through pro-oxidant antioxidant balance (PAB). The present study aimed to evaluate PAB and its association with high-sensitivity C-reactive protein (hs-CRP) in the serum of postmenopausal women with diabetes mellitus. METHODS: In this case-control study, 99 diabetic and 100 healthy postmenopausal women without diabetes mellitus were recruited. Serum PAB values, hs-CRP, lipid profile, insulin, and vitamin D levels were measured. Moreover, insulin resistance (HOMA-IR, HOMA-ß and QUICKI), waist circumference (WC), waist-to-hip ratio (WHR), waist-to-height ratio (WHtR), and body mass index (BMI) were calculated. RESULTS: Serum PAB, hs-CRP, insulin resistance, HOMA-ß, QUICKI, low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and triglycerides (TG) levels were significantly higher in the postmenopausal women with diabetes mellitus, while there was no significant difference in the total cholesterol (TC), serum insulin, WC, WHR, WHtR and vitamin D levels between the groups. Pearson correlation coefficient showed that HDL-C and insulin levels were directly correlated with serum PAB. Also, there was a significant direct relationship between LDL-C and insulin levels and hs-CRP. There was no meaningful relationship between serum insulin and vitamin D levels and other assessed parameters. Backward logistic regression showed a positive relationship between diabetes mellitus and serum PAB and an inverse relationship with serum HDL levels. CONCLUSIONS: Serum PAB, hs-CRP concentration, and lipid profile were significantly different between postmenopausal women with and without diabetes mellitus. These differences may contribute to the development of coronary complications.

Serum Prooxidant-Antioxidant Balance and hs-CRP in Patients with Clinical and Subclinical Hypothyroidism.

Background: Oxidative stress (OS) is caused by an imbalance between prooxidant substance production and antioxidant defense. OS is involved in physiologic interactions in the body and the pathogenesis of various disorders. This study aimed to evaluate serum prooxidant-antioxidant balance (PAB) as a selective prooxidant, antioxidant defense, and acute phase reactant protein in patients with subclinical and clinical hypothyroidism. Methods: This case-control study was conducted in three groups including clinical hypothyroidism (32 patients), subclinical hypothyroidism, (42 cases), and healthy controls (32 individuals). This study was performed in the Endocrine Clinic of Arash Training and Research Hospital, Tehran, 2017. In the study groups, thyroid hormones including T4 and Thyroid Stimulating Hormone (TSH), fasting blood glucose (FBG), lipid profile, PAB, and hs-CRP as inflammatory markers were measured and compared between the groups. Results: Among 106 participants, 95.3% were females, the gender balance was similar across groups and mean age was 30.79 ± 7.65 years. FBG and lipid profile except for cholesterol level were not significantly different between the three study groups. However, cholesterol level in the clinical hypothyroid group was significantly higher than the other two groups. PAB was higher in subclinical hypothyroidism compared to healthy controls after adjustment for age and TSH levels (P value: 0.04) but there was no significant difference in the clinical hypothyroid group in comparison with healthy controls. In addition, there was no significant difference in high-sensitivity C-reactive protein (hs-CRP) between the three study groups. Conclusions: This study suggests that that subclinical hypothyroidism increases PAB in comparison to healthy control which could indicate OS response in patients with subclinical hypothyroidism, respectively.

Prooxidant-antioxidant balance in stroke patients and 6-month prognosis.

BACKGROUND: Oxidative stress is thought to play an important role in the pathogenesis of stroke. METHODS: In the present study, we assessed the extent of oxidative stress in the sera of 120 patients in the first 24 hours after the onset of stroke and 80 healthy control subjects using a novel assay of prooxidant-antioxidant balance (PAB) that can measure the prooxidant burden and the antioxidant capacity simultaneously in one assay, thereby giving a redox index. RESULTS: In stroke patients median serum PAB values were significantly higher than that of the control group [125.50 (103.36 - 151.80) vs 43.40 (29.77 - 77.01), p < 0.001]. However, no significant difference was observed between subgroups of patients with transient ischemic attack (TIA), cerebral infarction or cerebral hemorrhage (p > 0.05). Median Serum PAB values were not significantly different in patients with different outcomes based on their 6-month prognosis (p > 0.05). CONCLUSIONS: We conclude that serum PAB values are elevated in patients with stroke, indicating a heightened state of oxidative stress, but do not distinguish between stroke types or predict 6-month prognosis.

Hypoxia-Induced miR-210 Overexpression Promotes the Differentiation of Human-Induced Pluripotent Stem Cells to Hepatocyte-Like Cells on Random Nanofiber Poly-L-Lactic Acid/Poly (ε-Caprolactone) Scaffolds.

An alternative treatment to liver transplantation includes the use of differentiated stem cells. Hypoxia has been shown to endow human-induced pluripotent stem cells (hiPSCs) with enhanced hepatic differentiation. We have investigated a new strategy for hepatocyte differentiation from hiPSCs using a three-step differentiation protocol with lentiviral overexpression of hypoxia-microRNA-210 of cells grown on a hybrid scaffold. We analyzed the transduction of the miR-210 lentiviral and definitive endoderm and pluripotency gene markers, including SRY-box 17 (SOX17), forkhead box A2 (FOXA2), and octamer-binding transcription factor 4 (OCT-4) by Real-Time PCR and fluorescent microscope. The scanning electron microscopy (SEM) examined the 3D cell morphological changes. Immunocytochemistry staining was used together with assays for aspartate aminotransferase, alanine aminotransferase, and urea secretion to analyze hepatocyte biomarkers and functional markers consisting of α-fetoprotein (AFP), low-density lipoprotein (LDL) uptake, fat accumulation, and glycogen. The flow cytometry analyzed the generation of reactive oxygen species (ROS). Compared to cells transfected with the blank lentiviral vectors as a control, overexpressing miR-210 was at higher levels in hiPSCs. The expression of endodermal genes and glycogen synthesis significantly increased in the differentiated lentiviral miR-210 cells without any differences regarding lipid storage level. Additionally, cells containing miR-210 showed a greater expression of ALB, LDL, AST, ALT, urea, and insignificant lower AFP and ROS levels after 18 days. However, SEM showed no significant differences between cells under the differentiation process and controls. In conclusion, the differentiation of hiPSCs to hepatocyte-like cells under hypoxia miR-210 may be a suitable method for cell therapy and regenerative medicine.

The association between daily naps and metabolic syndrome: Evidence from a population-based study in the Middle-East.

BACKGROUND: Daily naps are a common habit in many Middle Eastern and Asian countries; however, little is known about the association between daily naps and other health consequences, including the presence of metabolic syndrome (MetS). METHODS: Participants were recruited from the Mashhad stroke and heart atherosclerotic disorders study. We defined MetS according to International Diabetes Federation criteria. Nighttime sleeping hours were categorized into three categories: <6, 6-8, and >8 hours. Using logistic regression models, we analyzed the association between the duration of night-time sleep and daily naps with MetS and its different components. RESULTS: A total of 9652 individuals were included in the study: 3859 with MetS (40%) and 5793 without MetS (60%), as the control group. Of all, 72% participants had a regular daily nap. Those with daily naps had a higher odd of MetS [Odds ratio:1.19, confidence interval: (1.08-1.33); P < .001]. We also observed significantly higher odds of obesity, central obesity, hypertriglyceridemia, and diabetes or impaired fasting glucose in these subjects. Men sleeping <6 hours per night had a lower odd of MetS. However, we observed higher odds of cardiovascular risk factors in participants sleeping <6 hours, including obesity and diabetes or IFG. CONCLUSION: Napping is a common habit in middle Eastern countries. Although the cross-sectional design of our study cannot prove causality, we observed a significant association between the presence of MetS and daily naps. The public should be aware of this possibility and be educated about the importance of sleeping patterns.

Omega-3 fatty acid supplements improve the cardiovascular risk profile of subjects with metabolic syndrome, including markers of inflammation and auto-immunity.

OBJECTIVE: Fish-oil contains high concentrations of omega-3 fatty acids that have been shown to have anti-inflammatory properties. We have evaluated the effects of purified omega-3 fatty acid supplements on several anthropometric and biochemical parameters, including heat shock protein (Hsp) 27 antibody titres in subjects with metabolic syndrome. METHODS: Subjects (n = 120) with metabolic syndrome (mean age of 52.9 +/- 11.9 years) were randomly allocated to one of two groups: sixty subjects were given 1 gram of fish oil as a single capsule, containing 180 mg eicosapentaenoic acid and 120 mg docosahexaenoic acid daily for 6 months. Control subjects did not receive any supplementation over the same period. RESULTS: The study was completed by 47 subjects in the intervention group and 42 subjects in the control group. Treatment with omega 3 supplements was associated with a significant fall in body weight (P < 0.05), systolic blood pressures (P < 0.05), serum low density lipoprotein cholesterol (P < 0.05), and total cholesterol (P < 0.05), triglycerides (P < 0.05), high-sensitivity C-reactive protein (hs-CRP) (P < 0.01), and Hsp27 antibody titres (P < 0.05). No significant changes were observed in the control group. CONCLUSION: It appears that omega 3 improves the cardiovascular risk profile of subjects with metabolic syndrome, having effects on weight, systolic blood pressure, lipid profile and markers of inflammation and autoimmunity.

Three-dimensional nanofiberous PLLA/PCL scaffold improved biochemical and molecular markers hiPS cell-derived insulin-producing islet-like cells.

Nanofibrous scaffolds are considered as a new strategy for Type 1 diabetes mellitus therapy. We used a hybrid of poly-l-lactic acid (PLLA) and polycaprolactone (PCL) as three-dimensional (3D) culture models for differentiation of human induced pluripotent stem cells (hiPSCs) to beta islet-like cluster cell compared with routine culture (2D). Morphological changes of cells were checked by microscope. mRNA endodermal SOX-17 on day 7 and pancreatic gene markers Pdx1, glucagon and Glut2 were evaluated on day 23 by qPCR. As well as, insulin and C-peptide protein expression was evaluated by immunocytochemistry staining. In addition, insulin and C-peptide secretion in various glucose concentrations was evaluated by ELISA. Light and scanning electron microscopy (SEM) microscope showed changes in induced cells. In tandem, these modifications were more evident in 3 D culture. Pdx1, Glucagon and Glut2 markers in PLLA/PCL were significantly higher in 3 D culture. In addition, qualitative immunochemistry showed that insulin and C-peptide were expressed in 2 D and 3 D culture medium. Furthermore, evaluation of insulin and C-peptide clarified that secretion of these proteins in PLLA/PCL scaffold were statistically different in 2 D and 3 D strategies. These findings suggest that functional matured induction cells on PLLA/PCL scaffold can be used for islet beta cell therapy and regenerative medicine.

Expression Analysis of Previously Verified Fecal and Plasma Dow-regulated MicroRNAs (miR-4478, 1295-3p, 142-3p and 26a-5p), in FFPE Tissue Samples of CRC Patients.

BACKGROUND: Colorectal cancer (CRC) is one of the most common causes of cancer-related mortality worldwide. Early diagnosis of this neoplasm is critical and may reduce patients' mortality. MicroRNAs are small non-coding RNA molecules whose expression pattern can be altered in various diseases such as CRC. METHODS: In this study, we evaluated the expression levels of miR-142-3p, miR-26a-5p (their reduced expression in plasma samples of CRC patients was previously confirmed), miR-4478 and miR-1295-3p (their reduced expression in stool samples of CRC patients was previously confirmed) in tissue samples of CRC patients in comparison to healthy subjects. To achieve this purpose, total RNA including small RNA was extracted from 53 CRC and 35 normal subjects' Formalin-fixed, Paraffin-embedded (FFPE) tissue samples using the miRNeasy FFPE Mini Kit. The expression levels of these four selected miRNAs were measured using quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR). RESULTS: We found that the expression levels of miR-4478 and miR-1295b-3p (two previously down-regulated fecal miRNAs) were significantly decreased in FFPE samples of CRC patients compared to healthy controls. On the other hand, no significant differences were seen in expression levels of miR-142-3p and miR-26a-5p (two previously down-regulated circulating miRNAs) in FFPE samples between these two groups. CONCLUSION: Regarding current findings, it may be concluded that to diagnose CRC patients based on the miRNAs approach, stool samples are more likely preferable to plasma samples; nevertheless, additional studies with more samples are needed to confirm the results.

Microchips and their Significance in Isolation of Circulating Tumor Cells and Monitoring of Cancers.

In micro-fluid systems, fluids are injected into extremely narrow polymer channels in small amounts such as micro-, nano-, or pico-liter scales. These channels themselves are embedded on tiny chips. Various specialized structures in the chips including pumps, valves, and channels allow the chips to accept different types of fluids to be entered the channel and along with flowing through the channels, exert their effects in the framework of different reactions. The chips are generally crystal, silicon, or elastomer in texture. These highly organized structures are equipped with discharging channels through which products as well as wastes of the reactions are secreted out. A particular advantage regarding the use of fluids in micro-scales over macro-scales lies in the fact that these fluids are much better processed in the chips when they applied as micro-scales. When the laboratory is miniaturized as a microchip and solutions are injected on a micro-scale, this combination makes a specialized construction referred to as "lab-on-chip". Taken together, micro-fluids are among the novel technologies which further than declining the costs; enhancing the test repeatability, sensitivity, accuracy, and speed; are emerged as widespread technology in laboratory diagnosis. They can be utilized for monitoring a wide spectrum of biological disorders including different types of cancers. When these microchips are used for cancer monitoring, circulatory tumor cells play a fundamental role.

Formulation and Taste Masking of Ranitidine Orally Disintegrating Tablet.

Orally Disintegrating Tablets (ODT) have the advantages of both solid dosage form specially the stability and ease of handling and liquid dosage forms including ease of swallowing and pre-gastric absorption. We focused on taste masking and formulation of ranitidine ODT which disintegrates rapidly in the mouth within 60 sec using super-disintegrants, special polymers, water soluble and even insoluble excipients, sweeteners and essence. Various formulations were designed and made in four series. The amount of ranitidine in each formulation was 150 mg, and the final weight of tablets was around 500 mg. Prepared formulations were evaluated in terms of several physicochemical tests including powder/granule flowability, appearance, thickness, uniformity of weight, hardness, friability and disintegration time. Several taste masking techniques were investigated in each series of formulation, in order to cover the bitter taste of wranitidine. These included the addition of sweetener, granulation, solid dispersion with soluble and insoluble agents and complex formation with cellulose derivatives. The best formulation(s) in each group was/were chosen for taste evaluations with the help of 10 volunteers. Finally, formulation F14 was selected as the ultimate formulation, based on its better taste and shorter disintegration time (around 5 seconds). Formulation F14 contained Na CMC, avicel, Na starch glycolate, xylitol, saccharin, Na benzoate and menthol. The chosen formulation successfully passed the complementary evaluations such as assay of active ingredient and dissolution time. Na CMC was found to be acceptable in terms of decreasing disintegration time and enhanced taste masking potential and can be used in further ODT formulations.