فهرست مطالب عباس پیریایی

Renal fibrosis, with multiple aetiologies, is the typical pathological symptomof almost all chronic Kidney diseases and the most reliable indicator of CKD progression to end- stage renal failure. Kidney disease affects a variety of kidney cells and compartments and causes the spread of fibrosis called glomerulosclerosis, interstitial fibrosis, atherosclerosis, and perivascular fibrosis. Many cellular and molecular mechanisms are involved in the development of kidney fibrosis. Kidney disease affects a variety of existing cells and the compartment of the kidney and causes a variety of fibrosis called glomerulosclerosis, interstitial fibrosis, atherosclerosis, and perivascular fibrosis.

In this study, a comprehensive review of the cellular and molecular pathways involved in renal fibrosis, along with drug therapy and cell therapy used to improve the disease, by searching the databases of Scopus, PubMed, and Google Scholar was performed from 2010 to 2022.

Findings from studies showed that optimal cell therapy methods have fewer side effects than drug therapy. Due to the type of fibrosis involved in the kidney, it will be possible to repair it using stem cells. TGFB1 signaling seems to play an important role in the development of renal fibrosis, and drugs used to improve renal fibrosis often address the importance of the TGFB1 signaling pathway, which in addition to the relative improvement of the disease, has many side effects.: The use of cell therapy in the treatmentof chronic kidny fibrosis has more appropriate effects and fewer side effects than drug therapy, and mesenchymal stem cells are the most suitable candidates for transplantation.

Prostate cancer is the second leading cause of cancer deaths in men. Androgen ‎deprivation therapy has been identified to induce oxidative stress in prostate cancer, ‎leading to reactivation of androgen receptor (AR) signaling. Thus, antioxidant therapies ‎have gained attention as adjuvants for this cancer. Here, we report for the first time that ‎endostatin along with endometrial stem cells expressing TSP-1 anti-angiogenic gene ‎improved androgen- phenotype and genotype in rats prostate cancer cells.‎‏ ‏Forty male ‎rats (n=8) weighing approximately 95-100 g were purchased from Pasteur Inistitute of ‎Tehran . Animals were suffered to prostate cancer by injecting LNCaP cell line for 12 ‎weeks. Prostate cancer animals were immediately received endostatin for 28 days. ‎Prostate cancer animals were received stem cell containing the Tsp-1 anti-angiogenesis ‎gene. Histological examinations were measured. The levels of IL8 and VEGF serum ‎were assayed.‎‏ ‏In group receiving cells and drug, amount and height of acinus wall folds ‎decreased compared to the patient group and the shape of the vesicles was clear and the ‎epithelial lining were cohesive and regular. The levels of IL8 and VEGF serum were ‎significantly decreased in combination group than to prostate cancer group (P≤0.01). ‎

There is an urgent need for reliable in vitro liver models to assess the mechanisms of human diseases, and impact of newly developed drugs or toxins. Previously showed that co-culture of hepatocytes with non-parenchymal cells and presence of extracellular matrix (ECM) improve the in vitro functionality of hepatocytes. Aim of this study was to develop a reproducible method for scalable generation of liver microtissues.

In this experimental study, liver microtissues with 500um diameter generated through co-culturing of Huh-7, human mesenchymal stem cells and human umbilical vein endothelial cells in a composite hydrogel of liver-ECM and alginate. Cell viability, capability for glycogen storage and cytochrome P450 enzyme activity were evaluated on day 28. Hepatic-specific genes (ALB and SULT1A1) on days 1 and 14, and albumin and urea secretion on days 1, 7, 14, 21 and 28 were assessed. Furthermore, the microtissue sensitivity to acute toxicity of Metformin, Diclofenac and ethanol were investigated. Two-dimensional cultured Huh-7 was considered as control group.

Most cells in the microtissues were alive up to day 28, accumulated glycogen and have cytochrome inducibility 7-fold more than the basic activity. In comparison to control, ALB and SULT1A1 genes upregulated (p<0.05) on day 14, also albumin and urea secretion increased (p<0.05), in all sampling time, in microtissues. Furthermore, microtissues have more sensitivity (p<0.05) to some concentrations of the drugs and ethanol, compared to control.

It seems that the liver microtissues may considered as an appropriate and functional in vitro model for liver.

Hormonal disturbances in critical periods of fetal development can lead to changes in the physiology or morphology of an organ. The aim of the present study was to examine reproductive system in female rats exposed to a single dose of testosterone in one of their last fetal development days.

Testosterone (20 mg in 1 ml solvent) was subcutaneously injected to pregnant rats on gestational day 20. Control group received the solvent. Body weights of female offspring were measured at birth, at days of 15, 30, 45 and 60 of age and in adulthood, ano-genital distance (AGD) was determined at days of 6, 30 and 60 of age. Hormones and morphology of reproductive system and ovary tissue were examined in these offspring in adulthood. The comparison of two groups was performed using the Independent-sam ple student t-test.

In the offspring of experimental group, AGD, clitoris length, testosterone and LH concentrations were significantly increased but estradiol and anti-Mullerian hormone concentrations were significantly decreased (p<0.05). Moreover, the number of preantral and antral follicles was increased. However, the numbers of preovulatory follicles and corpora lutea were decreased but these changes were not significant statistically.

Exposure of female fetuses to a single dose of testosterone on day 20 of development leads to appearance of some developmental changes in external reproductive system and the appearance of disturbance in hormones secretion in adulthood.

Environmental factors and hormonal disorders during prenatal life can impair the evolution and differentiation of fetus’s reproductive system. The aim of this study was to the examination of sperm quality and testis tissue in adult rats that had exposed to exogenous testosterone during their prenatal life.
Pregnant rats divided into two experimental and control groups. In experimental group, pregnant rats daily were subcutaneously injected with 3 mg exogenous testosterone during gestational days 16-19, and controls received solvent. The reproductive system was examined in male offspring of these animals after puberty. Data analysis was performed by using the SPSS software (version 15). Distributions between groups are compared by using the Mann-Whitney test.
In the offspring of experimental group, motility and the number of sperm were significantly decreased (P The present study showed that prenatal exposure to testosterone leads to decrease of sperm quality and changes in testis tissue in male rats, after puberty.

Chronic wounds of diabetic patients, which resulted from some impairments including disruption in production and secretion of effective factors in the process of healing is highly regarded. In recent years, many studies have been reported the positive impact of low level laser therapy (LLLT) and mesenchymal stem cells (MSCs) and their conditioned media (CM) in diabetic wound healing. In the present study we evaluated the combined effect of LLLT and human-bone marrow (h-BM) MSCs-CM on the expression of an inflammatory factor and a growth factor, both involved in wound healing, in cultured fibroblasts at high glucose condition.
In this experimental study human dermal fibroblasts (HDFs) were cultured at culture media with normal and high glucose (as diabetic model) concentrations. Then the diabetic fibroblasts were treated by a laser beam with 0.5J/cm2 and h-BMMSCs-CM, separately and in combination with each other. Finally, the gene expression of bFGF and TGF-β2 were evaluated, using RT-PCR technique.
The expression of bFGF, significantly and the expression of TGF-β2, non-significantly were increased in all three treated diabetic groups, compared to the untreated diabetic group. Despite the relative expression of bFGF in diabetic group was 0.47±0.11. It significantly (p Apparently, by their synergistically effects, using the LLLT and the h-BMMSCs-CM together, positively affect the expression of tropic factor involved in diabetic wound healing.

Wound healing is a complex process that is impaired in diabetic patients due to several factors. So far, the positive effects of mesenchymal stem cells secretions in wound healing process have been reported. In this study, we investigated the effect of human mesenchymal stem cells Conditioned media on expression of effective factors involved in wound healing. 27 rats were divided into 5 groups: no wound control, normal control, diabetic control, diabetic placebo and diabetic experimental. Diabetes was induced by Alloxan. A wound was created on the back of the rats. Then, the conditioned medium was prepared from mesenchymal stem cells. Diabetic experimental rats received 200 microliter of conditioned medium intravenously. The wounds were sampled and expression of KGF and TGF-β1 genes was examined by RT-PCR on days four and seven after wounding. In the diabetic experimental group, expression of KGF gene at fourth and seventh days had been non-significantly increased in comparison to diabetic control group. While, expression of TGF-β1 gene in diabetic experimental group compared to diabetic control group had been significantly (p<0.05) increased on fourth day, and non-significantly increased on seventh day. It seems that using the conditioned medium derived from human mesenchymal stem cells positively affects the expression of trophic and inflammatory factors involved in diabetic skin wound healing.

The effects of non-invasive physical agents on wound and bone fracture healing have been considered by investigators of different medical fields. Several studies have been conducted about the effects of magnetic field on bone fracture healing. The purpose of this study was to investigate the effects of pulsed magnetic fields on healing of tibial bone lesion in rabbits. This experimental study was performed in Shahid Beheshti University of medical sciences in 2014. Eight New Zealand male rabbits were divided into control (with bone lesion) and experimental (with bone lesion and under pulsed magnetic field therapy) groups. A 4×10 cm bone lesion with 2 mm depth was made by surgery on both tibias of each rabbit. One month after surgery, the rabbits in the experimental group underwent 75 Hz pulsed magnetic field therapy with 4 mT intensity in lesion site for 30 days and 4 hours a day. Forty, 50 and 60 days after surgery, bone healing was evaluated with digital radiographs and bone density was measured in lesion sites. Data were analyzed using T- test. Mean relative bone density in lesion sites was 138.50, 150.73 and 168.30 for 8 bit grey scale in the experimental group during three consecutive imaging and showed %17.89, %15.46 and %16.59 increase in comparison with the control group. With regards to the results, it seems that pulsed magnetic field therapy improves healing of bone fracture through increasing bone density in fracture site and can be used after orthopedic surgeries.

Onosma dichroanthum Boiss. belongs to the Boraginaceae family and is one of the most important medicinal plants in north of Iran. This study aimed to examine the effect of the petroleum ether root extract ointment of Onosma dichroanthum Boiss. on healing of a surgically induced open skin wound in rats. In this study، 54 male adult Wistar rats were randomly allocated into control، vehicle، and experimental groups. A 20-mm cephalocaudal incision was made on the back skin of rats. Rats were sacrificed on days 4، 7 and 14 and histological examination (the number of fibroblasts، neutrophil، blood vessel sections and thickness of epiderm) was performed on skin samples. On day 4، the number of fibroblasts was significantly increased in the experimental group compared to the vehicle (P<0. 003) and control (P<0. 001) groups. On day 7، no significant difference was seen in the number of fibroblasts in the experimental group compared to the vehicle group (P<0. 680) and fibroblasts were significantly increased in the experimental and vehicle groups compared to the control (P<0. 001). On day 14، fibroblasts were significantly increased in the experimental and control groups compared to the vehicle (P<0. 001) and there was no significant difference in the number of fibroblasts in the experimental group compared to the control (P<0. 843). Also، no significant difference was seen in the number of neutrophils، blood vessel sections and thickness of epiderm on days 4، 7، 14 among the groups. Topical application of the petroleum ether root extract of onosma dichroanthum Boiss. has no significant effect on the healing of skin wound in rats.

Backgroundand

Production of a model of hepatic failure is used to evaluate the effectiveness and safety of alternative methods of liver transplantation. This study aimed to determine the appropriate dosage of carbon tetrachloride (CCl4) to induce acute hepatic failure in NMRI mice.

In this study, at first, a mixture of CCl4 dissolved in olive oil was administered intraperitoneally to 5 groups of 6 mice. After 24 hours, serum level of liver enzymes and the status of liver tissue were evaluated. To investigate the survival of mice, CCl4 (at doses of 0.5, 1, 1.5, 2, and 2.5ml/kg.bw) was administered to 6 new groups of mice, and these animals were monitored for 4 days. To assess the effect of the toxin in the next days, the determined dose was administered to 24 new mice, and serum and histological evaluations were repeated.

The serum level of liver enzymes and the degree of hepatic lesions were significantly increased with increased dose of CCl4. Values more than 1ml/kg of CCl4 induced acute liver damage. Also, the value of 1ml/kg decreased mortality rate and values more than 2ml/kg increased mortality rate. Maximum increase in serum levels of liver enzymes was observed 24 hours after injection of CCl4 (at the dose of 1.5ml/kg), which decreased gradually. The degree of liver damage in days after the injection was the same until the fourth day, but the liver regeneration phasewas initiated after 72 hours.

According to the results of this study, a single intraperitoneal administration of CCl4 at the dose of 1.5mL/kg CCl4 could be an appropriate dosage for the production of a model of acute hepatic failure in NMRI mouse.

Diabetes is a metabolic chronic disorder that makes it difficult to heal wounds because of its complications. To this point, many studies on the use of mesenchymal stem cells in wound healing process particularly diabetic ulcers have been done. In this study, we evaluate the impact of mesenchymal stem cells-derived conditioned medium (MSC-CM) on diabetic wound strength. The type of study is experimental. 24 rats were divided into four groups: normal control, diabetic control, diabetic placebo (culture medium treated) and diabetic experimental (treated with MSC-CM). Diabetes mellitus was induced in the diabetic rat groups by intraperitoneal injections of alloxan. One full-thickness skin incision was made on the dorsal region of each rat.Conditioned medium were obtained from human bone marrow mesenchymal stem cells. 24 hours after wounding, diabetic experimental rats received 200 µl of conditioned medium, and diabetic placebo rats received 200 µl of of culture medium intravenously. 15 days after wounding, the wounds were sampled and biomechanical testing of elasticity (tensiometery) was performed on them and strength of wounds was evaluated by ANOVA test. Body weight and blood glucose difference of diabetic rats was significantly higher than the control group (P=0.00). Maximum force of experimental group was 3.45±1.05 while this criterion in diabetic control was 1.76± 0.56 and in diabetic placebo was 2.30±0.62, which was significantly increased (p<0.05). Morover,the maximum stress of experimental group was 0.34±0.1, which compared to the diabetic control group (0.18±0.05) showed a significant increase. In addition, standard elastic stiffness of the experimental group was 3.19 ±1.07, whichcompared with diabetic placebo group (2.2 ±0.19) significantly increased. It seems to be human mesenchymal stem cells-derived conditioned medium has significant effect on wound strength in diabetic rats.

Onosma dichroanthum Boiss. is one of the most important mountainous medicinal plants in Iran. This study was done to determine the biomechanical evaluation of root extract ointment of Onosma dichroanthum Boiss. on wound healing in rats. In this experimental study, 18 male adult Wistar rats were randomly allocated into control (I), vehicle (II) and treated group with ointment containing of extract 1% of root of Onosma dichroanthum Boiss., (III). 20 mm vertical skin incision wound were made on rats back side. The assessment of the wound healing was carried out at day 14. At the end of study, rats were sacrificed, skin sample were extracted and evaluated by biomechanical method (maximum force, elastic stiffness, energy absorption). There was no significant difference in biomechanical parameters among the treated, vehicle and control groups. Topical application of Onosma dichroanthum Boiss. root have no effect on healing of skin wound in animal model.

Nitric Oxide (NO) is a signaling molecule produced by the NO Synthases (NOS) enzime and is known to incorporate in the regulation of many reproductive events that occur in the Fallopian tube (FT). The aim of this study was to investigate the production of endothelial NOS (eNOS) in the ciliated and secretory cells of luminal epithelium of FT and alteration of eNOS during parturition. In this case-control study a total of 20 FTs samples obtained from two groups of women including: 10 FTs from the women at luteal phase of the menstrual cycle and 10 FTs of healthy pregnant women during parturition. Samples were fixed in 10% buffered formalin and paraffin sectioned for immunohistochemical evaluation. Localization of eNOS in the ciliated and secretory cells of luminal epithelium of FT was seen in both groups. However، we did not found anyexpression of eNOS in smooth muscle cells of all groups (p>0. 05). No significant difference in expression of eNOS was seen between luteal phase and healthy pregnant groups. Regarding the no significant differences in expression of eNOS in ciliated and secretory cells of luminal epithelium between pregnant women at term and women at luteal phase of menstrual cycle، the results suggested that activity and function of cilia in both groups are same.

The aim of this study was to evaluate the expression of eNOS in the FTs of women with tEP. In this case-control study, a total number of 30FTs samples were obtained from three groups including: 10 FTs of women that bearing an EP, 10 FTs from the non-pregnant women at luteal phase of the menstrual cycle, and 10 FTs of healthy pregnant women (n=10). Samples were fixed in 10% buffered formalin and then were evaluated by immunohistochemistry. Localization of eNOS was seen in secretory and ciliated luminal epithelium and vascular endothelium of all groups. However, we did not observed the expression of eNOS in smooth muscle cells of all groups. Expression of eNOS in luminal epithelium of women with EP compared to non-pregnant women at luteal phase of menstrual cycle and healthy pregnant group showed statistically significant increase (p=0.00). Significant difference in expression of eNOS was not observed in luminal epithelium of FTs of women at luteal phase compared to healthy pregnant groups (p=0.78). This study indicates that changes in expression of eNOS in luminal epithelium of FT may lead to development of EP.

Wound healing represents a dynamic physiological process influenced by many factors. The aim of the present study was to evaluate the effects of hydro-alcoholic extracts of aerial components of Teucrium polium on the tensile strength in rat. control groups. pressure ulcer were made over the dorsal thoracic region according to the shyn model. Animals were treated with topical hydro-alcoholic extracts of aerial components of Teucrium polium twice a day post surgery until complete healing was achieved. Tensiometry were then studied. Twelve Wistar rats were randomly divided into tow equal (n= 6) treatment and No significant difference in biomechanical parameters was observed among the ointment containing of extract 1%، vehicle and control groups. polium did not have any effect on healing of pressure ulcer in an animal model. Further studies are requiring for assessment of other extracts by different solvents of this plant on pressure ulcer wound healing. This study shows that topical application of Teucrium polium. Aerial components of Teucrium.

Pentoxifylline decreased blood viscosity، improved peripheral blood circulation، and improved red blood cell flexibility. It also induced sperm motility and increased microvascular blood circulation in non-diabetic humans. The aim of present study was to evaluate the effect of pentoxifylline administration on Sertoli and Leydig cells count of experimentally induced type 1 diabetes in male rats. In this experimental study 50 male rats were randomly divided into 3 non-diabetic groups، and 2 diabetic groups. Type 1 diabetes was induced by injection of streptozotocin، and rats were held for 30 days. Experimental groups on non-diabetic and diabetic rats were received pentoxifylline daily. One non-diabetic group received daily normal saline. Then، all rats were anesthetized and their right testis were extracted and were examined histologically. Sertoli cells and Leydig cells were counted. Data were analyzed by ANOVA and LSD tests. Increase in number of pentoxifylline- treated diabetic rats cells (7. 43±0. 86، 15. 25±1. 6) were significantly higher than control diabetic rats (4. 4±0. 38، 11±1. 31) (p=0. 001 and p=0. 000 respectively). There were significant difference between pentoxifylline treated diabetic rats cells (7. 43±0. 86، 15. 25±1. 6)، and normal saline- treated non- diabetic rats cells (7. 17±1. 13، 25. 8، 3. 5) with other groups (p=0. 001 and p=0. 000 respectively). There were no significant difference between non- diabetic groups. Our results showed that pentoxifylline administration increased Sertoli cells and Leydig cells count in non-diabetic and diabetic rats in compare to control groups. However its effect in diabetic rats were significant.

Spinal cord has a limited capacity to repair; therefore، medical interventions are necessary for treatment of injuries. Transplantation of Schwann cells has shown a great promising result for spinal cord injury (SCI). However، harvesting Schwann cell has been limited due to donor morbidity and limited expansion capacity. Furthermore، accessible sources such as bone marrow stem cells have drawn attentions to themselves. Therefore، this study was designed to evaluate the effect of bone marrow-derived Schwann cell on functional recovery in adult rats after injury. Mesenchymal stem cells were cultured from adult rats’ bone marrow and induced into Schwann cells in vitro. Differentiation was confirmed by immunocytochemistry and RT-PCR. Next، Schwann cells were seeded into collagen scaffolds and engrafted in 3 mm lateral hemisection defects. For 8 weeks، motor and sensory improvements were assessed by open field locomotor scale، narrow beam، and tail flick tests. Afterwards، lesioned spinal cord was evaluated by conventional histology and immunohistochemistry. In vitro observations showed that differentiated cells had Schwann cell morphology and markers. In this study، we had four groups (n = 10 each): laminectomy، control، scaffold and scaffold + Schwann cells. Locomotor and sensory scores of cell grafted group were significantly better than control and scaffold groups. In histology، axonal regeneration and remyelination were better than control and scaffold groups. This study demonstrates that bone marrow-derived Schwann cells can be considered as a cell source for Schwann cells in SCI treatment.

Porpose:To evaluate the development of cardiomyocyte-like cells derived from human bone marrow mesenchymal stem cells induced by 5-azacytidine (5-aza) under in vitro condition. Human bone marrow mesenchymal stem cells (BMMSCs) were purified, then the cells were induced by 5µM 5-aza for 5 weeks. To transdifferentiate into cardiomyocyte, the culture medium of human BMMSCs was changed every 24 hours, in the way that the cells were put in the culture medium containing 5-aza for 24 hours and, then were put in a culture medium free of the said material for another 24 hours, and this process continued until sampling. RT-PCR assay was performed to detect the expression of specific myocardium genes including α-actinin, myosin heavy chain (MHC) and connexin-43 each weeks after first induction. Fluocytometry was used to determine the expression of specific myocardium proteins including α-actinin, troponin and connexin-43 at 3rd and 5th weeks after the first induction. The expression of all three myocardium genes was detected 1 to 5 weeks after induction in a time-dependent manner. There was a continuous increase in expression of α-actinin and connexin-43 genes from the 1st to the 5th week, while mRNA of MHC gene was at the highest level in the forth week. The expression of α-actinin and troponin proteins was equal after three and five weeks, whereas that of the connexin-43 was higher after five weeks in comparison to the 3rd week. The current study indicated that stimulating human bone marrow mesenchymal stem cells by 5-aza with the concentration of 5µM under in vitro condition can lead to differentiating cardiomyocyte-like cells. Furthermore prolonged culture duration may lead to more differentiated cells

Background and Application of low-frequency stimulation (LFS) induces anticonvulsant effects. In this study, the effect of changes in LFS frequencies on its anticonvulsant effects in kindling model of epilepsy was investigated by determining the behavioral and ultrastructural changes. Methods and Materials: In this experimental study, 45 Wistar rats weighing 250-280 g were used. For induction of kindled seizures, stimulating and recording electrodes were implanted in perforant path and dentate gyrus respectively. Animals were stimulated in a rapid kindling manner. Different groups of animals received LFS at different frequencies (0.5, 1 and 5 Hz) following kindling stimulations and their effects on kindling rate were determined using behavioral and electrophysiological studies. After stimulating the animals for 7 days, they were killed and their dentate gyri were sampled for ultrastructural studies under electron microscopy. For data analysis, one-way and two-way ANOVA, LSD, Kruskal Wallis and Mann Whitney U were used in Statistica 2. Different LFS frequencies had a significant inhibitory effect on kindling rate and decreased after-discharge duration and the number of stimulations to achieve stage 4 and 5 seizures significantly. In addition, application of LFS prevented the increase in the post-synaptic density and induction of concave synaptic vesicles following kindling. There was not any significant change between anticonvulsant effects of LFS at different frequencies. Obtained results show that LFS application can prevent the neuronal hyper-excitability by preventing the ultrastructural changes during kindling, and can exert its anticonvulsant effects.

To study the impact of ultraweb nano-fibrillar substrates on in vitro differentiation of mouse bone marrow-derived mesenchymal stem cells into hepatocyte-like cells.

For a duration of up to 36 days, mouse bone marrow mesenchymal stem cells (MSCs) were cultured on an artificial basement membrane containing ultraweb nanofiber (nano+) and plates (nano-) coated with Matrigel and used growth and differentiating factors for hepatogenic differentiation. Then, to study the impact of nanofibers on hepatocyte differentiation, mRNA expressions for liver-specific genes (CK7, AFP, FOXa2, ALB, CYP1a1, HNF4a) were analyzed using real-time PCR, and ultrastructures of differentiated cells were evaluated using scanning and transmission electron microscope.

Both experimental groups expressed mRNA of liver-specific genes in a time dependent manner. However, mRNA expression trends of liver-specific genes, especially those of FOXa2 and HNF4a, were better in the nano+ culture as compared to the nano- culture (p<0.05). Furthermore, the ultrastructures of differentiated cells in the 36 day-old nano+ culture were more mature as compared to those of the nano- group.

The results suggest that topographical properties of the extracellular matrices produced by ultraweb nanofibers are effective for the in vitro reproduction of more differentiated MSC-derived hepatocyte-like cells.

The aim of the present study was to isolate mesenchymal stem cells (MSCs) from canine bone marrow, differentiate them into bone and examine the structure of the produced bone that was not reported before. MSCs from canine bone marrow were extracted, expanded and cultured in an osteogenic medium which differentiated into bone tissue on the surface of the cover slips. At the end of the differentiation period, the bone prepared was examined with light and electron microscopes. In this study, the isolated cells were also differentiated into adipocytic cell lineage. Cultivated mesenchymal stem cells formed several nodule-like structures during differentiation period. According to the light microscopic images, these nodules were composed of several layers of cells with an abundant matrix among the cells. Transmission electron microscopic study indicated that the cells were mainly of spindle-shape morphologly similar to osteocytic cells, and the matrix consisted of collagen fibers with sharp striation organized in bundles perpendicular to each other. It seems that the bone produced from in vitro differentiation of the MSCs, histologically is similar to that of laminar bone particularly in terms of the collagen fiber arrangement.

Background and Vitro cartilage differentiation of mesenchymal stem cells (MSCs) has been noticed in several investigations. In this regard, almost always molecular differentiation of the cells has been examined, while structural and morphological differentiation of them has been ignored. Therefore, the present study examines the structure and ultrastructure of the cartilage differentiated from murine MSCs compared with that of costal cartilage. 2× 105 MSCs isolated from the bone marrow of NMRI mice were pleted by centrifugation and cultured for 21 days in chondrogenic medium. At the end of cultivation period, occurrence of chondrogenic differentiation was confirmed by reverse transcriptase–polymerase chain reaction (RT-PCR) analysis for some cartilage-specific genes. To compare the structure of differentiated tissue with that of natural cartilage, the cartilage was differentiated from MSCs and the cartilage obtained from the same murine rib was prepared for transmission electron microscopy (TEM). Structural studies indicated that similar to the costal cartilage, the cartilage produced from differentiation of perichondrium-like layer was formed. According to the microscopic images, in contrast to costal chondrocytes, the differentiated cells had euchromatic nucleus and their cytoplasm contained plenty of the organelles involved in active cell secretion. Furthermore, intercellular matrix in differentiated cartilage had a fibrillar appearance. Our results indicated that the structure of cartilage produced in micro mass culture system is somewhat different from that of costal cartilage. The cells from differentiated tissue seemed to be more active than those from costal cartilage.

Background and ObjectivesMesenchymal stem cells are appropriate candidates to treat diseases including articular cartilage defects. There are plenty of researches being conducted to make the application of these cells possible. The purpose of this study was to cultivate murine mesenchymal stem cells (MSCs) in alginate gel and transplant them subcutaneously to immuno-suppressed rats to examine their chondrogenic potential in vivo.Materials and Methods 4-6 week old NMRI mice were sacrificed and their bone marrow cells were cultivated in 6-cell plates at the density of 500 cell/cm2. The pure fibroblastic cells appeared after two passages. 2×106 fibroblastic cells were mixed with 1 ml of alginate solution and converted into gel by being exposed to calcium chloride solution. MSCs-embedded alginate gel were then transplanted subcutaneously to 6 rats that had received an immunosuppressive drug (cyclosporine) for transplant rejection to be avoided. 5 weeks after transplantation, the alginate gels were removed and evaluated by histochemistry, RT-PCR for certain cartilage markers, and transmission electron microscopy.Results5 weeks after transplantation, the skin was incised and the alginate gel with its surrounding vascular connective tissue were removed. Tuloidine blue staining indicates that the cells within the gel assumed oval morphology and occupied lacuna-like cavities. RT-PCR analysis revealed that in these cells the mRNA of some cartilage markers such as collagen II (the marker of hyaline cartilage), collagen X (hypertrophied chondrocyte marker in osteogenesis), and aggreacan were largely produced. Ultra-thin sections analysis showed that the cells within the lacuna-like cavity of alginate gel contain a large amount of expanded rough endoplasmic reticulum and secret fibrillar extra cellular matrix. ConclusionsTransplanted murine MSCs cultivated in alginate gel can differentiate into hyaline cartilage with the sign of osteogenic initiation.