The Impact of Calcium Hydroxide on the Osteoinductive Capacity of Demineralized Freeze-Dried Bone Allograft: an In-Vitro Study.

STATEMENT OF THE PROBLEM: A great challenge in periodontal therapy is the regeneration enhancement of osseous defects through applying osteoinductive materials. Demineralized freeze-dried bone allograft (DFDBA) has already been introduced as an allograft with osteoconductive and variable osteoinductive properties. Calcium hydroxide [Ca(OH)2] is an available well-known material in dentistry, which induces hard tissue formation. PURPOSE: This study evaluated the efficiency of combination of DFDBA and Ca(OH)2 in improving the quality of osteoinduction of DFDBA. MATERIALS AND METHOD: Human bone marrow-derived mesenchymal stem cells were taken from volunteers' iliac crest. Cell proliferation was determined by MTT test at 18, 24 and 48 hours post-culture in 10 groups. The employed material were 0.5, 1.0 mg/ml Ca(OH)2 in two forms of suspension and pH-adjusted solution, 10mg/ml DFDBA per se and in combination with 0.5 and 1.0 mg/ml Ca(OH)2. Mineralization was assessed by Alizarin red staining in 10 mg/ml DFDBA, DFDBA+ 0.5 and 1 mg/ml Ca(OH)2 in solution and suspension forms. The data were statistically analyzed by using one-way ANOVA and Tukey's post-hoc test (p< 0.05). RESULTS: The pH-adjusted solutions exhibited better cell proliferation compared with the suspension groups. The combination of 0.5mg/ml Ca(OH)2 solution and DFDBA increased the cell proliferation and mineralization compared with DFDBA per se (p= 0.033). CONCLUSION: The combination of Ca(OH)2 with DFDBA improved the osteoinductivity of DFDBA.

Comparison of healing intra-articular fracture of distal femur using a Kirschner wire and autologous fibrin glue in an animal model.

Considering different surgical techniques for the fixation of osteochondral intra-articular fracture, the present study aimed to compare the efficacy of autologous fibrin glue and Kirschner wire (KW) on an osteochondral fracture in the left lateral condyle of Dutch rabbits with a control group. After 6 weeks, macroscopic and microscopic evaluation showed that autologous fibrin glue is easier and faster with a higher number of bone trabecula (P<0.05), whereas the healing rate and cellularity of the healing site were not different between the two groups (KW and glue). The use of autologous fibrin glue can be an alternative to KW fixation in the fixation of osteochondral fractures. LEVEL OF EVIDENCE: Level II.

Design and Fabrication of a Novel Transplant Combined with Human Bone Marrow Mesenchymal Stem Cells and Platelet-rich Fibrin: New Horizons for Periodontal Tissue Regeneration after Dental Trauma.

Avulsed teeth that are replanted dried are more prone to loss. Recent tissue engineering studies focus on fabrication of various cell delivery systems to improve the overall prognosis of such teeth. To evaluate this new cell transplant method, we initially aimed at designing of PRF scaffold and determining BMMSCs viability and function on the fabricated scaffold. To test this concept in-vitro, human BMMSCs were isolated and characterized by cell surface marker, and their potential on osteogenic/adipogenic differentiation. The biological effects of PRF scaffold on human BMMSCs were then investigated by cell proliferation assay. The data were quantified for statistical analysis. It was found that PRF significantly induced BMMSCs proliferation throughout the incubation period due to its growth factor secretion. Results of MTT assay showed an increasing trend during the testing period from day 1 to day 7. The result of scanning electron microscopy showed that BMMSCs could tightly adhere to fibrin scaffold just shortly after seeding. These data suggest that the BMMSCs/PRF construct has the potential to improve the clinical prognosis of replanted avulsed teeth in future. Additional studies are needed to be performed before its clinical use.

Selective toxicity of persian gulf sea cucumber holothuria parva on human chronic lymphocytic leukemia b lymphocytes by direct mitochondrial targeting.

Natural products isolated from marine environment are well known for their pharmacodynamic potential in diversity of disease treatments such as cancer or inflammatory conditions. Sea cucumbers are one of the marine animals of the phylum Echinoderm. Many studies have shown that the sea cucumber contains antioxidants and anti-cancer compounds. Chronic lymphocytic leukemia (CLL) is a disease characterized by the relentless accumulation of CD5+ B lymphocytes. CLL is the most common leukemia in adults, about 25-30% of all leukemias. In this study B lymphocytes and their mitochondria (cancerous and non-cancerous) were obtained from peripheral blood of human subjects and B lymphocyte cytotoxicity assay, and caspase 3 activation along with mitochondrial upstream events of apoptosis signaling including reactive oxygen species (ROS) production, collapse of mitochondrial membrane potential (MMP) and mitochondrial swelling were determined following the addition of Holothuria parva extract to both cancerous and non-cancerous B lymphocytes and their mitochondria. Our in vitro finding showed that mitochondrial ROS formation, MMP collapse, and mitochondrial swelling and cytochrome c release were significantly (P < 0.05) increased after addition of different concentrations of H. parva only in cancerous BUT NOT normal non-cancerous mitochondria. Consistently, different concentrations of H. parva significantly (P < 0.05) increased cytotoxicity and caspase 3 activation only in cancerous BUT NOT normal non-cancerous B lymphocytes. These results showed that H. parva methanolic extract has a selective mitochondria mediated apoptotic effect on chronic lymphocytic leukemia B lymphocytes hence may be promising in the future anticancer drug development for treatment of CLL. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1158-1169, 2017.

Evaluation of bone marrow derived mesenchymal stem cells for full-thickness wound healing in comparison to tissue engineered chitosan scaffold in rabbit.

BACKGROUND: Chronic wounds present a major challenge in modern medicine. Even under optimal conditions, the healing process may lead to scarring and fibrosis. The ability of mesenchymal stem cells (MSCs) to differentiate into other cell types makes these cells an attractive therapeutic tool for cell transplantation. Both tissue-engineered construct and MSC therapy are among the current wound healing procedures and potential care. Chitosan has been widely applied in tissue engineering because of its biocompatibility and biodegradability. AIM: The aim of the current work was to compare the efficiency of MSCs and chitosan dressing, alone or in combination treatment on wound healing. METHODS: This study was conducted on 15 rabbits, which were randomly divided in 3 groups based on the type of treatment with MSCs, chitosan dressing and combination of both. A full-thickness skin defect was excised from the right and left side of the back of each animals. Defects on right sides were filled with treatments and left side defects were left as control. Evaluation of the therapeutic effectiveness was performed through a variety of clinical and microscopical evaluations and measurements of the process of wound healing on days 7, 14, 21, and 28. Histological evaluation of wound healing was classified by different scoring systems. RESULTS: The data indicated that wounds treated with bone marrow derived MSC had enhanced cellularity and better epidermal regeneration. During the early stages of wound healing, the closure rate of bone marrow derived MSC-treated wounds were significantly higher than other treatments (P<0.05). Although the MSCs in the wound edges enhance the healing of the full-thickness wound, the healing process of chitosan treatment was slower than the control group. CONCLUSION: This study revealed advanced granulation tissue formation and epithelialization in wounds treated with MSCs, and may suggests this treatment as an effective applicant in wound healing process. Chitosan scaffold dressings, whether alone or in combination with MSCs, have worsened the wound healing as compared to the control group.

A case-report of delayed repositioning of intruded permanent maxillary central incisors accompanied by complicated crown fractures: A 2-year follow-up.

Intrusive luxation is the most severe type of dental injury with a complex healing sequence. Pulp necrosis, root resorption (surface, inflammatory and replacement resorption), and defects in marginal periodontal bone healing are the main complications. Treatment strategies can be either active, by repositioning (surgical or orthodontic extrusion), or passive, by spontaneous re-eruption based on the thorough evaluation of the case. This paper reports a case of delayed repositioning of severely intruded permanent maxillary central incisors accompanied by complicated crown fractures after 3 months. After thorough clinical and radiographic evaluations, and based on guidelines, the teeth were surgically repositioned and splinted for 6 weeks. One week after the initial intervention, the endodontic treatment for both permanent maxillary incisors were initiated using calcium hydroxide. 6 months later, the teeth were ready for MTA plug and gutta-percha root canal filling. During the follow-up period, the teeth had remained functional and esthetically acceptable. Further yearly observations are planned at least for 5 years.

Growth suppression effect of human mesenchymal stem cells from bone marrow, adipose tissue, and Wharton's jelly of umbilical cord on PBMCs.

OBJECTIVES: Immunosuppressive property of mesenchymal stem cells (MSCs) has great attraction in regenerative medicine especially when dealing with tissue damage involving immune reactions. The most attractive tissue sources of MSCs used in clinical applications are bone marrow (BM), adipose tissue (AT), and Wharton's jelly (WJ) of human umbilical cord. The current study has compared immunomodulatory properties of human BM, AT, and WJ-MSCs. MATERIALS AND METHODS: Three different types of human MSCs were isolated, cultured, and characterized by flow cytometry and differentiation potentials. The MSCs were co-cultured with allogeneic phytohemagglutinin (PHA) activated peripheral blood mononuclear cells (PBMCs). The proliferation of PBMCs was assessed by flow cytometry of carboxyfluorescein succinimidyl ester (CFSE) stained cells and compared to each other and to the growth of PBMCs in the absence of MSCs. Additionally, the growth suppression was indirectly assessed by using the transwell culture system. RESULTS: The proliferation of PBMCs reduced to 6.2, 7 and 15.4- fold in cultures with AT-MSCs, WJ-MSCs, and BM-MSCs, respectively, compared to the PHA-activated cells. When the growth suppression was indirectly assessed by using the transwell culture system, it was revealed that AT-MSCs, WJ-MSCs, and BM-MSCs caused growth reduction in PBMCs to 3, 8, and 8 -fold, respectively, compared to the PHA-activated cells. CONCLUSION: These data collectively conclude that the immunomodulatory effects of MSCs, which may mostly carry out through direct cell to cell contact, are different between various sources. Accordingly results of this study may contribute to the application of these cells in cell therapy and regenerative medicine.

An in vitro model for hepatocyte-like cell differentiation from Wharton's jelly derived-mesenchymal stem cells by cell-base aggregates.

AIM: The present study investigated the differentiation potential of human Umbilical Cord Mesenchymal Stem Cells (UCMSCs) into hepatic lineage through embryonic body-like aggregate formation in the presence of IGF-1. BACKGROUND: Cells derived from Wharton's jelly have been reported to display a wide multilineage differentiation potential, showing some similarities to both embryonic (ESC) and mesenchymal stem cells (MSCs). PATIENTS AND METHODS: Human MSCs isolated from the umbilical cord were plated in 20 µL micro drops. A two-step differentiation protocol was used and the cell aggregates were exposed to the media supplemented with IGF, HGF, oncostatin M, and dexamethasone for 21 days. Immunoperoxidase and immuno-fluorescence were performed for cyrokeratins 18, 19 and albumin. Functional assays were done by periodic acid Schiff (PAS) and indocyanine green. RESULTS: The expression of cytokeratin 19 was shown to be higher in the cells derived from 3D spheroids compared to those cultured in conventional protocol. They showed a polygonal shape after being exposed to hepatogenic media. Immunostaining demonstrated the expression of cytokeratin-18, 19 and albumin by the differentiated cells. Besides, PAS staining revealed glycogen storage in differentiated cells. Also, a greater number of large size differentiated cells were found at the periphery of the expanded cell aggregates. CONCLUSION: We established a protocol for UCMSC differentiation into hepatocytes and these cells were morphologically and functionally similar to hepatocytes. Thus, hepatocyte differentiation may be facilitated by the UCMSCs aggregate formation before administration of the differentiation protocols.

Wharton's Jelly-derived Mesenchymal Stem Cells can Differentiate into Hepatocyte-like Cells by HepG2 Cell Line Extract.

BACKGROUND: Wharton's jelly is an unlimited source of stem cells that can be used in cell therapy and tissue engineering without any ethical concern. It has been revealed the cell-free extract could be effective to induce cell differentiation. The objective of this study was to induce Wharton's jelly-derived mesenchymal stem cells (MSCs) into hepatocyte-like cells by premeabilization of the cells in the presence of HepG2 cell line extract. METHODS: MSCs were isolated from the umbilical cord, CD marker profile and their differentiation potential into adipogenic and osteogenic lineages were determined. The cells were then, permeabilized by streptolysin O in the presence of HepG cell extract. The treated cells were cultured for 17 days. The cell phenotype was evaluated and the hepatocyte specific markers were detected by immunofluorescence and immunocytochemistry. The Periodic Acid Schiff (PAS) reaction and the cellular uptake of indocyanine green were performed to evaluate the functional behavior of the differentiated cells. RESULTS: The phenotype of extract-treated MSCs changed into a round or polygonal cells with few short processes and they could express high level of albumin, cytokeratin 18 and 19. The MSCs also could store glycogen and uptake and release indocyanine green. CONCLUSION: We demonstrated for the first time that Wharton's jelly-derived MSCs could differentiate into hepatocyte-like cells by premeabilization of them in the presence of HepG2 cell extract. This study suggests a feasible method to differentiate MSCs into functional hepatocyte-like cells.

Effect of Platelet-Rich Plasma on CCl4-Induced Chronic Liver Injury in Male Rats.

Platelet-rich plasma (PRP) has been of great concern to the scientists and doctors who are involved in wound healing and regenerative medicine which focuses on repairing and replacing damaged cells and tissues. Growth factors of platelet-rich plasma are cost-effective, available, and is more stable than recombinant human growth factors. Given these valuable properties, we decided to assess the effect of PRP on CCl4-induced hepatotoxicity on rats. The rats received CCl4 (1 mL/kg, i.p. 1 : 1 in olive oil) twice per week for 8 weeks. Five weeks after CCl4 injection, the rats also received PRP (0.5 mL/kg, s.c.) two days a week for three weeks. Twenty-four hours after last CCl4 injection, the animals bled and their livers dissected for biochemical and histopathological studies. Blood analysis was performed to evaluate enzyme activity. The results showed that PRP itself was not toxic for liver and could protect the liver from CCl4-induced histological damages and attenuated oxidative stress by increase in glutathione content and decrease in lipid peroxidative marker of liver tissue. The results of the present study lend support to our beliefs in hepatoprotective effects of PRP.

Involvement of TNF-α in differential gene expression pattern of CXCR4 on human marrow-derived mesenchymal stem cells.

Cell therapy and tissue repair are used in a variety of diseases including tissue and organ transplantation, autoimmune diseases and cancers. Now mesenchymal stem cells (MSCs) are an attractive and promising source for cell-based therapy according to their individual characteristics. Soluble factors which are able to induce MSCs migration have a vital role in cell engraftment and tissue regeneration. Tumor necrosis factor α (TNF-α) is a major cytokine present in damaged tissues. We have investigated the pattern of gene expression of chemokine receptor CXCR4 in nine groups of human bone marrow-derived MSCs stimulated with TNF-α in different dose and time manner. Comparison of TNF-α treated with untreated MSCs revealed the highest expression level of CXCR4 after treatment with 1, and 10 ng/ml of TNF-α in 24 h, and the production of CXCR4 mRNA was regulated up to 216 and 512 fold, respectively. Our results demonstrated the differential gene expression pattern of chemokine receptor CXCR4 in human marrow-derived MSCs stimulated with inflammatory cytokine TNF-α. These findings suggest that in vitro control of both dose and time factors may be important in stem cell migration capacity, and perhaps in future-stem cell transplantation therapies.

Major barriers responsible for malnutrition in hemodialysis patients: challenges to optimal nutrition.

BACKGROUND: Nutritional barriers may contribute to malnutrition in hemodialysis (HD) patients. Higher rates of morbidity and mortality rates have been reported in malnourished HD patients. These patients are faced with different challenges affecting their nutritional status. OBJECTIVES: The aim of this cross-sectional study was to identify most important barriers responsible for malnutrition in HD patients. PATIENTS AND METHODS: We randomly selected 255 of 800 stable HD patients from three HD centers with an age range of 18-85 years, who had been on hemodialysis for at least three months without any acute illness. Each patient was interviewed to evaluate malnutrition [subjective global assessment (SGA), malnutrition inflammation score (MIS)], and potential medical, behavioral and socioeconomic barriers. Body composition of patients was checked through bioelectrical impedance analysis (BIA). Routine clinical markers of malnutrition such as serum albumin and total protein were measured using standard automated techniques. Binary logistic regression model was used to find the association between nutritional markers and potential barriers. RESULTS: Patients with higher SGA had lower knowledge about general nutrition [odds ratio (OR), 1.3], potassium (OR, 1.89), difficulty chewing (OR, 1.16), and shopping (OR, 1.16). Those with greater MIS scores had poor appetite (OR, 1.3), depression (OR, 1.21), and difficulty with cooking (OR, 1.15). Lower BCM (body cell mass) was associated with poor appetite (OR, 0.92) and needed help for cooking (OR, 0.88). Patients with higher BFMI (body fat mass index) had insufficient general nutrition (OR, 1.15), and protein (OR, 1.27) knowledge, and needed help for shopping (OR, 1.14). Moreover, patients with higher SGA scores were those with older age and longer duration of HD. CONCLUSIONS: Three medical barriers (poor appetite, depression and difficulty chewing), one behavioral barrier (poor total nutrition, protein, and potassium knowledge), and one socioeconomic barrier (needing help for shopping and cooking) were independently associated with nutritional markers.

Adiponectin as a novel indicator of malnutrition and inflammation in hemodialysis patients.

INTRODUCTION: Protein-energy malnutrition and inflammation are common and overlapping conditions in hemodialysis patients, which are associated with increased risk of morbidity and mortality. Adiponectin is an adipocytokine exclusively produced by adipose tissue. The aim of this study was to further elucidate the association between serum adiponectin levels and the nutritional status of hemodialysis patients. MATERIALS AND METHODS: Seventy-three patients on hemodialysis for at least 3 months, three times weekly, without any acute illness, were divided into 2 groups of well-nourished (n = 25) and malnourished (n = 48) based on their nutritional status, measured by the subjective global assessment. Serum levels of adiponectin, albumin, blood urea nitrogen, and creatinine; body mass index; and the malnutrition-inflammation score were measured in all patients. These values were compared between well-nourished and malnourished patients. The correlations of nutritional variables with serum levels of adiponectin were determined, as well. RESULTS: Except for the malnutrition-inflammation score, which was significantly higher in the malnourished patients compared to the well-nourished ones (11.1 +/- 3.6 versus 4.2 +/- 2.0, P < .001), no other significant differences were found between the two groups. A weak but significant positive correlation was found between the serum levels of adiponectin and subjective global assessment scores (r = 0.25, P = .03). CONCLUSIONS: The results of our study point to potential utility of serum adiponectin level as an indicator of nutritional status in hemodialysis patients. Further studies are needed to clarify the role of adiponectin in the pathogenesis of malnutrition in hemodialysis patients.

Selenium supplementation improves the nutritional status of hemodialysis patients: a randomized, double-blind, placebo-controlled trial.

BACKGROUND: Malnutrition is highly prevalent in hemodialysis (HD) patients. These patients have high levels of oxidative stress and inflammation which can subsequently induce malnutrition. Selenium levels have been found to be decreased in HD patients. As selenium deficiency leads to oxidative stress and inflammatory response, the aim of this study was to evaluate the effects of selenium supplementation on oxidative and inflammatory markers and the nutritional status of HD patients. METHODS: In this randomized double-blind placebo-controlled trial, 80 patients on stable HD for at least 3 months without any acute illness or active infections were randomly allocated to two equal groups to receive one selenium (200 µg) or placebo capsule daily for 12 weeks. Serum levels of lipoproteins, malondialdehyde (MDA), interleukin-6 (IL-6), high-sensitivity C-reactive protein (HSCRP), homocysteine, ferritin and transferrin as well as the subjective global assessment (SGA) score, malnutrition-inflammation score (MIS) and hemoglobin (Hb) levels were measured at the baseline and at the end of the treatment phase. The primary outcome was a change in the nutritional status measured by the SGA score from the baseline towards the end of the treatment phase of the study. RESULTS: The SGA score and MIS decreased significantly in the selenium group compared to the placebo group (P < 0.001 for both). Moreover, serum levels of MDA decreased significantly in the selenium group compared with increasing levels in the placebo group (P < 0.001). Selenium supplementation also hindered an increase in IL-6 levels compared with the placebo group (P = 0.016). There were no significant differences between the selenium and placebo groups in terms of changes in serum levels of lipoproteins, HSCRP, homocysteine, ferritin and transferrin or Hb levels. CONCLUSIONS: This study shows that selenium may be an effective complementary supplement for reducing the severity of malnutrition in HD patients through alleviating oxidative stress and inflammation.

Vitamin D receptor genotype in pancreas allograft: a pilot study.

OBJECTIVES: Transplanting of pancreatic grafts is an established treatment for diabetes mellitus. Polymorphisms in genes, coding for proteins involved in an immune response, may influence immunologic and nonimmunologic mechanisms that lead to allograft loss. Vitamin D receptor agonists have been shown to increase long-term allograft survival in humans. MATERIALS AND METHODS: Twenty-one pancreatic recipients transplanted in the Transplantation center of Shiraz University of Medical Sciences were selected and genotyped for the polymorphism of the vitamin D receptor genes (FokI), and the association of each genotype with acute rejection was evaluated. A control group of 100 unrelated otherwise healthy individuals, from the Iranian Blood Transfusion Organization were enrolled. The individuals were selected from Shiraz (a city located in Southern Iran), and the genotype frequency was compared with control group. RESULTS: The overall prevalence acute rejection was 28% (6/21). In the genotype study, homozygous FF presented in 15 patients (71%), heterozygous Ff presented in 6 patients (29%), and no homozygous ff was identified. In the control group, there were 50% with FF, 48% with Ff, and 2% with the ff genotype identified. The only genotype that was detected in rejection group was FF, while the frequency of FF in the nonrejection group was 60%. CONCLUSIONS: This study examined several patients to determine whether the vitamin D receptor (FokI) genotype is involved in acute allograft rejection and requires deeper investigation.

Conditions to improve expansion of human mesenchymal stem cells based on rat samples.

AIM: To improve the isolation and expansion of human marrow-derived mesenchymal stem cells (MSCs) based on rat samples. METHODS: Based on the fact that rat MSCs are relatively easy to obtain from a small aspirate, bone marrow-derived MSCs from rat were cultured and characterized to set up the different protocols used in this study. Then, accordingly, almost the same protocols were performed on human healthy bone marrow samples, after obtaining approval of the ethics committee and gaining informed consent. We used different protocols and culture conditions, including the type of basal media and the culture composition. The MSCs were characterized by immunophenotyping and differentiation. RESULTS: There was no difference in morphology and proliferation capacity between different culture media at the first passage. During the 5-7th passages, the cells gradually lost their morphology and proliferation potential on Dulbecco's modified Eagle's medium (DMEM) high glucose and α modified Eagle's medium. Although the cells expanded rapidly for up to 10 passages on DMEM low glucose containing 10% to 15% fetal calf serum (FCS), their proliferation was arrested without change in morphology and differentiation capacity at the third passage on 5% FCS. Flow cytometric analysis and functional tests confirmed that more than 90% of marrow cells which were isolated and expanded by our selective protocols were MSCs. CONCLUSION: We improved the isolation and expansion of human bone marrow derived MSCs, based on rat sample experiments, for further experimental and clinical use.

Insulin-like growth factor 1 (IGF-I) improves hepatic differentiation of human bone marrow-derived mesenchymal stem cells.

The ability of MSCs (mesenchymal stem cells) to differentiate between other cell types makes these cells an attractive therapeutic tool for cell transplantation. This project was designed to improve transdifferentiation of human MSCs into liver cells using IGF-I (insulin-like growth factor 1) which, despite its important role in liver development, has not been used for in vitro hepatic differentiation. In the present study, the MSCs derived from healthy human bone marrow samples were cultured and characterized by immunophenotyping and differentiation potential into osteoblast and adipocytes. Transdifferentiation into hepatocyte-like cells was performed in the presence/absence of IGF-I in combination with predefined hepatic differentiation cocktail. To evaluate transdifferentiation, morphological features, immuno-cytochemical staining of specific biological markers and hepatic functions were assessed. Morphological assessment and evaluation of glycogen content, albumin and AFP (α-feto protein) expression as well as albumin and urea secretion revealed statistically significant difference between experimental groups compared with the control. Morphology and function (albumin secretion) of IGF-I-treated cells were significantly better than IGF-I-free experimental group. To the best of our knowledge, our study is the first to demonstrate that the combination of IGF-I with the predefined hepatic differentiation cocktail will significantly improve the morphological features of the differentiated cells and albumin secretion.

Hepatogenic differentiation of mesenchymal stem cells induced by insulin like growth factor-I.

AIM: To improve hepatic differentiation of human mesenchymal stem cell (MSC) using insulin growth factor 1 (IGF-I), which has important role in liver development, hepatocyte differentiation and function. METHODS: Bone marrow of healthy donors was aspirated from the iliac crest. The adherent cells expanded rapidly and were maintained with periodic passages until a relatively homogeneous population was established. The identification of these cells was carried out by immunophenotype analysis and differentiation potential into osteocytes and adipocytes. To effectively induce hepatic differentiation, we designed a protocol based on a combination of IGF-I and liver specific factors (hepatocyte growth factor, oncostatin M and dexamethasone). Morphological features, hepatic functions and cytological staining were assessed to evaluate transdifferentiation of human marrow-derived MSCs. RESULTS: Flow cytometric analysis and the differentiation potential into osteoblasts and adipocytes showed that more than 90% of human MSCs which were isolated and expanded were positive by specific markers and functional tests. Morphological assessment and evaluation of glycogen storage, albumin and α-feto protein expression, as well as albumin and urea secretion revealed a statistically significant difference between the experimental groups and control. CONCLUSION: In vitro differentiated MSCs using IGF-I were able to display advanced liver metabolic functions, supporting the possibility of developing them as potential alternatives to primary hepatocytes.

Value of serum cystatin C as a marker of renal function in the early post kidney transplant period.

Management of renal transplant patients requires periodic measurement of renal function especially in early post transplant period. This is usually assessed by measuring the creatinine clearance, but because of its limitations, it is not an ideal marker for assessing the renal function. Serum cystatin C (sCyC) appears to be an endogenous marker of glomerular filtration rate (GFR). To assess the use of sCyC as a marker of renal function in kidney transplant patients, we compared it with serum creatinine (sCr) and 24-hour urine creatinine clearance (CrCl) in the first week post-transplantation. Among 60 patients (62.8% men, 37.2% women) undergoing kidney transplantation (average age: 44.87 +/- 13.37 years), we determined renal function at 1, 3, 5, and 7 days after kidney transplantation using: sCr, sCyC and CrCl in a 24-hours urine specimen. During the first 5 days following transplantation, there was a progressive decline in sCr levels. In the first 5 days, post transplantation we could not find good correlation between CrC and sCyC, and the sCyC increased during these 5 days, but after that in day 7, there was a good correlation between CrC and sCyC which is coinciding with decreasing the dose of steroid (r= .625). Therefore, we recommend using sCyC may be used as a marker of renal function after one-week post kidney transplantation.

A fast and easy nitroblue tetrazolium method for carrier screening and prenatal detection of chronic granulomatous disease.

BACKGROUND: Analysis of the functional activity of neutrophils is of great importance in the differential diagnosis of patients with recurrent bacterial infections. It has long been established that stimulated polymorphonuclear leukocytes reduce nitroblue tetrazolium. Application of a simple and reliable nitroblue tetrazolium method that clearly differentiates the chronic granulomatous disease patients and heterozygote carriers in some groups suspected to have chronic granulomatous disease was investigated. METHODS: This study consisted of 197 samples taken from 100 children (2 - 24-month-old) and 81 neonates (aged < 2 months) referred to our center either due to a suspected bacterial infection or suspected immunodeficiency. The sample also included 16 cord blood samples. Fifty healthy adult individuals were enrolled in this study and were diagnosed as normal control. Neutrophil reduction of nitroblue tetrazolium can be stimulated in vitro by protein kinase agonists such as phorbol myristate acetate, resulting in release of superoxide anion. RESULTS: Phorbol myristate acetate is an exceptionally powerful stimulant and when used in conjunction with glass adherence, caused nearly all normal neutrophils to become transformed and reduced nitroblue tetrazolium to formazan deposits. Of 197 blood samples, 9 were diagnosed as having unrelated chronic granulomatous disease and 7 were carriers of X-linked or autosomal recessive chronic granulomatous disease. The carriers had a range of 15 - 75% stimulated neutrophils. CONCLUSION: We have established a phorbol myristate acetate-stimulated nitroblue tetrazolium test for detection of chronic granulomatous disease patients, which clearly differentiates the chronic granulomatous disease patients from heterozygote carriers. The results in cord fetal blood indicate that this test may be used for antenatal diagnosis of affected boys, carrier females, and autosomal recessive variants of chronic granulomatous disease. The technique is simple, fast, inexpensive, and requires only a few microliters of blood.