This study was aimed to investigate whether the thrombopoietin (rhTPO) may facilitate myelofibrosis or not. The modified Dexter culture system with various concentrations of rhTPO was used to culture the stromal cells in vitro; the proliferative activity of cells was detected by MTT method; the morphologic changes were observed by light and scanning electron microscopy; the staining changes of ALP, PAS, AS-D NCE and IV type collagen were observed by cytochemistry method; the changes of fibronectin, laminin and IV type collagen were assayed by immunohistochemistry method; the cell surface antigens were assayed by flow cytometry. The results indicated that rhTPO could promote the proliferation of stromal cells which was related to the concentrations of rhTPO. Proliferative activity of stromal cells increased with increasing of rhTPO concentration, and was not related to the exposure time. On day 3 stromal cells adhered to the wall, and became oval. On day 7 stromal cells turned to fusiform and scattered dispersively. On day 12 to 14 these cells ranged cyclically and became long fusiform. Cells covered 70%-80% area of bottle bottom at that time. By day 16 to 18 these cells covered more than 90% area of bottom and ranged cyclically. They displayed the same shape as fibroblasts. By light microscopy with Wrights-Giemsa staining, fibroblasts predominated morphologically, few macrophages, endothelial cells and adipose cells were found. There were no significant differences between experimental group and control group. On day 14 to 42 the adherent cells were positive with PAS staining, poorly positive with ALP and naphthol AS-D chloroacetate esterase (AS-D NCE) staining, and the difference in cytochemistry was not significant between two groups. When these cells were dyed with Masson's trichrome and Gomori's staining, neither collagen fibers nor reticular fibers were positive, but fibronectin, laminin, and collagen type IV appeared positive stronger in experimental group than those in control. The expressions of these molecules were not dependent on culture time. By scanning electron microscopy microvilli and fibers on cell surface appeared more and more, monolayer cells evolved into multilayer cells, and newly-formed fibroblasts appeared gradually as culture time prolonged. These alterations were not different among various groups. The expressions of CD34, CD45, CD105, CD106, and CD166 were not affected obviously by rhTPO. It is concluded that rhTPO had no effects on histochemical properties of stromal cells. Fiber staining and scanning electron microscopic examinations revealed that rhTPO can not facilitate fiber formation of stromal cells. But rhTPO may be able to augment the expressions of fibronectin, laminin and collagen type IV of stromal cells. Therefore it is still necessary to follow up the patients for a long time, who have received rhTPO therapy clinically.
Bone Marrow Cells ; cytology ; drug effects ; Cell Proliferation ; drug effects ; Fibroblasts ; Humans ; Stromal Cells ; cytology ; drug effects ; Thrombopoietin ; pharmacology

OBJECTIVETo investigate SiO2-induced EMT in human bronchial epithelial cells HBE in vitro.

METHODSHBE cells were cultured and then stimulated with indicated doses of SiO2 (0, 50, 100, 200, 300 µg/ml). The morphological changes were observed by microscope. In addition, Western blot was per-formed to detect the expression of E-cad, α-SMA and Vim. The changes of migration ability were examined by wound-healing assay in vitro.

RESULTS(1) After exposure to SiO2, HBE cells lost contact with their neighbor and displayed a spindle-shape, fibroblast-like morphology. (2) Compared with the control, the E-cad (300 µg/ml group) expression downregulated 2.98 fold (P < 0.05), and the Vim (300 µg/ml group) and α-SMA (200 µg/ml group) expression upregulated 4.46 fold and 3.55 fold (P < 0.05). There were significant differences between 100, 200, 300 µg/ml groups and the control group (P < 0.05). (3) In the test group, the percentage of wound-healing areas/wound areas were larger than those in control group (P < 0.05).

CONCLUSIONSSiO2 could induce EMT in human bronchial epithelial cells.

Bronchi ; cytology ; Cells, Cultured ; Epithelial Cells ; cytology ; drug effects ; Epithelial-Mesenchymal Transition ; drug effects ; Humans ; Silicon Dioxide ; adverse effects ; Stromal Cells ; cytology ; drug effects

Bone marrow stromal cells (BMSCs), a kind of stem cells residing in bone marrow, have self-renewal, high proliferative capacity and the potential of multilineage differentiation. It has a good prospect in application of the cell replacement therapy, the gene therapy and the tissue engineering and so on. As the content of BMSCs is extremely low in bone marrow, BM-SCs must be amplified in vitro and induced to differentiation to meet the clinical needs. Researches of the recent years suggest there is a very promising way that Chinese medicine could induce BMSCs proliferation, differentiation. Based on the Chinese medicine theory, "the kidney generating marrow and dominating bone" and "kidney storing essence, essence and marrow", the TCM scholars have done some researches to explore the function of warming yang and reinforcing kidney of Chinese medicine to promote bone marrow stromal cells and found that these drugs can promote the BMSCs to proliferate and to differentiate into osteogenic, cartilage and nerve cells. This article elaborates and presents the researches on this aspect.
Animals ; Bone Marrow Cells ; cytology ; Cell Differentiation ; drug effects ; Humans ; Medicine, Chinese Traditional ; Stromal Cells ; cytology

OBJECTIVETo investigate the effect of ecdysterone (EDS) on the proliferation of human bone marrow mesenchymal stem cells (hMSCs) in vitro.

METHODShMSCs were isolated from human bone marrow cell suspension by density gradient centrifugation. The expression of integrins CD44, CD105, CD34 and CD29 were examined by immunocytochemical method. EDS at 10, 25, 50 or 100 microg/ml were added in hMSC culture system, using the routine culture medium for hMSCs as control. The cell viability were analyzed by MTT assay and the cell cycle changes were examined by flow cytometry.

RESULTSThe optical density (OD) differed significant between the EDS treatment groups and the control group (P<0.01), and 25 microg/ml EDS group showed the highest OD value (P<0.01) without significant differences among 10, 50 and 100 microg/ml EDS groups (P>0.05). Flow cytometry showed that treatment of the cells with 25 microg/ml EDS significantly increased the cell percentages in S and G(2)M phases and the proliferation index (PI) of the cells as compared with the control group.

CONCLUSIONWithin a given concentration range, EDS can promote the proliferation of hMSCs in vitro, and this effect can be the most obvious at the concentration of 25 microg/ml. The effect of EDS in promoting the proliferation of hMSCs does not positively correlate to EDS concentration administered.

Adult ; Cell Proliferation ; drug effects ; Cells, Cultured ; Ecdysterone ; pharmacology ; Humans ; Male ; Mesenchymal Stromal Cells ; cytology

OBJECTIVETo investigate the role of mesenchymal stem cells (BMSCs) in multiple myeloma (MM) bone marrow (BM) microenrivonment and their effect on myeloma cells survival and bortezomib induced apoptosis.

METHODSBMSCs were derived from BM of untreated myeloma patients (MM-BMSCs) and healthy donors (HD-BMSCs), respectively. The phenotype, proliferation time and cytokine secretion of MM-BMSCs were detected and compared with HD-BMSCs. Then BMSCs were co-cultured with myeloma cell line NCI-H929 and bortezomib in vitro. The NCI-H929 cells proliferation and bortezomib induced cell apoptosis were investigated.

RESULTSMM-BMSCs and HD-BMSCs were isolated successfully. The phenotype of MM-BMSCs was similar to that of HD-BMSCs. Expressions of CD73, CD105, CD44 and CD29 were positive, but those of CD31, CD34, CD45 and HLA-DR (< 1%) negative. The proliferation time of MM-BMSCs was longer than that of HD-BMSCs (82 h vs 62 h, P < 0.05). Moreover, over-expressions of IL-6 and VEGF in MM-BMSCs culture supernatant were detected as compared with that in HD-BMSCs [(188.8 ± 9.4) pg/ml vs (115.0 ± 15.1) pg/ml and (1497.2 ± 39.7) pg/ml vs (1329.0 ± 21.1) pg/ml, respectively]. MM- BMSCs supported survival of the myeloma cells NCI-H929 and protected them from bortezomib induced cell apoptosis.

CONCLUSIONSMM-BMSCs is benefit for myeloma cells proliferation and against cell apoptosis induced by bortezomib. Over-expression of IL-6 and VEGF maybe play a critical role in these effects.

Apoptosis ; drug effects ; Bone Marrow Cells ; cytology ; Bortezomib ; Humans ; Mesenchymal Stromal Cells ; metabolism ; Multiple Myeloma ; metabolism

We aimed to examine the effect of pioglitazone on transdifferentiation of preosteoblasts from rat bone marrow mesenchymal stem cells (BMSCs) into adipocytes and investigate its effect on bone metabolism. BMSCs were harvested from the femurs and tibias of a rat, then separated, purified, proliferated for 3 generations and differentiated into preosteoblasts for 5 days and 14 days respectively in the presence of osteogenic medium. Thereafter, the preosteoblasts were cultured for 21 days in the presence of adipogenic medium with and without pioglitazone (1 μg/mL). Partially-differentiated osteoblasts were identified by mineralized nodules with Alizarin red S staining. Transdifferentiated adipocytes were identified by Oil Red O staining. Reverse transcription PCR (RT-PCR) was performed to assay the expression levels of osteogenic markers Runx2 and ALP, and an adipogenic marker PPARγ. Those cells cultured for 5 days did not show mineralized nodules as detected by staining of Alizarin red S, while those cultured for 14 days showed dispersed mineralized centers in the form of brown spots, although without obvious red mineralized nodules. After adipogenic transdifferentiation for 21 days, adipose-drops were found in cells of 5CG and 5EG earlier than those of 14CG and 14EG, and the former showed much more adipocytes separately as detected by Oil Red O staining. Whatever the time was 5 days or 14 days of BMSCs osteogenic differentiation, the cells cultured with pioglitazone showed much more adipocytes than those without pioglitazone. Our experiment showed that the less time it took for BMSCs osteogenic differentiation, a stronger ability remained for BMSCs to transdifferentiate into adipocytes. The mRNA expression levels of Runx2 and ALP were decreased by 1.79 and 1.90 times respectively in 5EG (P< 0.05) as compared with 5CG, and that of PPARγ was increased by 1.31 times in 5EG (P<0.05) as compared with 5CG. The mRNA expression levels of Runx2 and ALP were decreased by 1.45 and 1.54 times respectively in 14EG (P<0.05) as compared with 14CG, and that of PPARγ was increased by 1.39 times in 14EG (P<0.05) as compared with 14CG. It was concluded that pioglitazone stimulated the transdifferentiation of BMSCs into adipocytes. These observations provided a potential mechanism of imbalance in thiazolidinedione induced bone metabolism.
Adipocytes ; drug effects ; Animals ; Cell Transdifferentiation ; drug effects ; Female ; Male ; Mesenchymal Stromal Cells ; drug effects ; Osteoblasts ; drug effects ; Osteogenesis ; drug effects ; Rats ; Rats, Sprague-Dawley ; Thiazolidinediones ; pharmacology

Bone marrow mesenchymal stem cells (MSCs) have active abilities of self-replication and multidifferentiation. In recent years, a lot of studies have proved that MSCs can be induced and differentiated into nerve-like cells under certain conditions. Because of some advaced characteristics including sampling convenience, no immune rejection, high transfection rate and stable exogenous gene expression, MSCs will provide new way in treating disease of nervous system. In this article, the research progress of bone marrow mesenchymal stem cells differentiation into nerve-like cells induced by Traditional Chinese Medicine shall be discussed, and explore the research thinking guided by basis theory of TCM.
Animals ; Bone Marrow Cells ; cytology ; drug effects ; Cell Differentiation ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Mesenchymal Stromal Cells ; cytology ; drug effects ; Neurons ; cytology ; drug effects

Bone marrow mesenchymal stem cells (BMSCs) are a kind of pluripotent stem cells derived from bone marrows, which can not only support hematopoiesis, but also have capabilities of multidifferentiation, high-proliferation and self-renewing. They have become one of hotspots in stem cell studies. Studies on in vitro intervention with BMSCs with TCMs have made remarkable progress in recent years. According to the findings, some traditional Chinese medicines can promote proliferation of BMSCs, some can inhibit the apoptosis of BMSCs, while others can induce BMSCs to differentiate into multiple cell types, such as osteoblast. Furthermore, some studies also involved relevant action mechanisms. The authors summarized the advance in relevant studies by reference to relevant literatures of this field.
Animals ; Apoptosis ; drug effects ; Bone Marrow Cells ; cytology ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Humans ; Medicine, Chinese Traditional ; methods ; Mesenchymal Stromal Cells ; cytology ; drug effects

Bone marrow mesenchymal stem cells (BMSC) are a group of cells with the capability of self-renewal and potential of multilineage differentiation. It was reported that some traditional Chinese medicines and their extract have the potential to induce BMSC differentiate into multiple cells. For example, notoginse nosides can induce BMSC differentiate into neurons and myocytes, naringin can induce BMSC differentiate into osteoblast, youguiyin-containing rabbit serum can induce BMSC differentiate into chondrocyte. This article reviews some progresses of this area in recent years.
Animals ; Bone Marrow Cells ; cytology ; drug effects ; Cell Differentiation ; drug effects ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Mesenchymal Stromal Cells ; cytology ; drug effects

The purpose of this study was to investigate the effect of salidroside on human bone marrow mesenchymal stem cell (hBMMSC) apoptosis induced by cytarabine C (Ara-C) and its mechanism, hBMMSC were cultured in vitro and isolated by Fircoll density gradient centrifugation; cell surface antigens were measured by flow cytometry; the osteogenic and adipogenic differentiation of MSC was tested and evaluated by specific staining methods. The proliferation and apoptosis of cells exposed to Ara- C were detected by MTT and flow cytometry respectively. The experiments were divided into 4 groups: control group, Ara-C group, salidroside group and Ara-C+salidroside group. The mRNA expression of BCL-2 and BAX was assayed by RT-PCR. The results showed that the adherent cells displayed spindle and fibroblast cell-like shape; the hBMMSC expressed CD44, CD71 and HLA-ABC, not expressed CD34, CD45 and HLA-DR; the hBMMSC successfully differentiated into osteogenic and adipogenic lineages, which showed mineralization with von Kossa staining. Furthermore, liquid vacuoles were detected by oil red O staining; Ara- C exhibited a less inhibitory effect on the proliferation of hBMMSC treated with salidroside. The apoptosis of hBMMSC treated with salidroside were significantly higher as compared with control group (P < 0.05); RT-PCR results demonstrated that the BCL-2 expression was significantly down regulated but BAX mRNA expressions was up-regulated in Ara- C group as compared with those in the control group. Salidroside significantly inhibited the apoptosis of MSC and reversed the mRNA expression of BCL-2 and BAX. It is concluded that salidroside can inhibit the apoptosis of hBMMSC induced by Ara-C, its mechanism may be related with the regulation of BCL-2/BAX expression.
Apoptosis ; drug effects ; Bone Marrow Cells ; cytology ; drug effects ; Cells, Cultured ; Cytarabine ; pharmacology ; Glucosides ; pharmacology ; Humans ; Mesenchymal Stromal Cells ; cytology ; drug effects ; Phenols ; pharmacology