Xiaoguang Chen

A new method for Schwann-like cell differentiation of adipose derived stem cells.

Peripheral nerve repair can be enhanced by Schwann cell transplantation, but the clinical application of this procedure is limited by donor site morbidity and the inability to quickly generate a sufficient number of cells. Thus, alternative cell systems for the generation of Schwann cells are desired. Schwann-like cell induced from adipose-derived stem cells (ADSCs) may be one of the ideal alternative cell systems for Schwann cell generation. Although co-culture with Schwann cells or chemicals combined with a mixture of glial growth factors are often utilized for Schwann cell-like differentiation of ADSCs, these methods are usually complicated or expensive. In this experiment, the rat sciatic nerve was cut, and then soaked in culture medium for two days. The treated culture medium was used as an induction agent after filtering. The obtained ADSCs were incubated with the above induction culture medium for five days. Then, expression of the typical Schwann cell markers, S-100 and GFAP proteins was determined by immunocytochemical staining and Western blotting. The results showed that almost all of the treated ADSCs displayed a spindle shape like morphology after being incubated with induction culture medium for 24h and expressed S-100 and GFAP proteins after five days. All of these characteristics of differentiated rat ADSCs were similar to genuine Schwann cells. Thus, this new method, which utilized trophic factors secreted from sciatic nerve leachate, was capable of inducing ADSC differentiation into Schwann-like cell.

Adipose-derived stem cells undergo spontaneous osteogenic differentiation in vitro when passaged serially or seeded at low density

Adipose-derived stem cells (ADSCs) are a convenient source of cells for regenerating tissue. Widespread application of ADSCs requires that they propagate efficiently and differentiate in vitro. We investigated the differentiation potential of ADSCs during long-term expansion in vitro and when the cells were seeded at low density. ADSCs were isolated from the inguinal fat pads of 3-week-old male rats, then cultured serially for 12 passages; some ADSCs at passage 3 were seeded at low density. The differentiation potential of ADSCs from passage 3 to passage 12 was assessed by their capacity for adipogenesis and osteogenesis while cultured in specific induction media. Spontaneous osteogenesis of ADSCs at passage 12 and of ADSCs that were seeded at low density was detected by western blotting, alizarin red S staining and measurement of alkaline phosphatase (ALP) activity. We found that with increasing passage number, the adipogenic potential of ADSCs decreased and osteogenic differentiation increased. Alizarin red S staining, bone morphogenetic protein-2 (BMP-2) and runt-related transcription factor 2 (Runx2) expressions, and ALP activity demonstrated that both ADSCs at passage 12 and those that were seeded at low density differentiated into osteoblasts without additional induction factors.

Silencing of phospholipase C gamma 2 promotes proliferation of rat hepatocytes in vitro

The management of hepatic failure is undoubtedly difficult, and poor results have led to the search for novel therapeutic approaches. Nowadays, anti‐apoptotic gene therapy is considered as an ideal approach. It has been proved that phospholipase Cγ2 (PLCγ2) is involved in the apoptosis of immune cells and tumor cells; however, whether this gene is related to hepatocyte death is still unclear. This study examined the role of PLCγ2 by inhibiting its expression in rat hepatocytes with siRNA. We also further analyzed the cellular mechanism by which the expression inhibition of PLCγ2 induces cell death. Silencing PLCγ2 gene by adenovirus vector expressing PLCγ2‐targeted siRNA caused the great decline in the number of G1‐ and G2/M phase cells, the significant increase in the number of S phase cells, and the obvious reduction in apoptosis index. In addition, silencing PLCγ2 gene relieved the rat hepatocyte damage, such as the cell shrinkage and chromatin condensation, nuclear fragmentation. Further analysis of Ad‐PLCγ2 siRNA‐transfected hepatocytes demonstrated that suppression of PLCγ2 gene expression could cause the caspase dependent cell death by inhibiting the signal pathway MEKK1/MKK4/JNK1/2/c‐Jun. In conclusion, these findings suggest that interference with PLCγ2 expression could relieve the inhibitory effect of PLCγ2 on hepaocyte apoptosis, thus, promote proliferation through inactivating PKCδ‐mediated JNK1/2 signaling pathway.

PLCγ2 promotes apoptosis while inhibits proliferation in rat hepatocytes through PKCD/JNK MAPK and PKCD/p38 MAPK signalling

The PLCG2 (PLCγ2) gene is a member of PLC gene family encoding transmembrane signalling enzymes involved in various biological processes including cell proliferation and apoptosis. Our earlier study indicated that PLCγ2 may be involved in the termination of regeneration of the liver which is mainly composed of hepatocytes, but its exact biological function and molecular mechanism in liver regeneration termination remains unclear. This study aims to examine the role of PLCγ2 in the growth of hepatocytes.

Enhancement or inhibition of PLCγ2 expression in rat hepatocytes by recombinant adenoviral vectors that contain full-length gene or siRNA

Abstract We investigated the effects of recombinant adenovirus vectors that overexpress or silence PLCγ2 on the expression of this gene during hepatocyte proliferation. Hepatocytes were isolated, identified by immunofluorescent cytochemical staining and infected by previously constructed Ad-PLCγ2 and Ad-PLCγ2 siRNA1, siRNA2 and siRNA3. Green fluorescent protein (GFP) expression was observed by fluorescence microscopy. Infection percentage was calculated by flow cytometry. mRNA and protein levels of PLCγ2 were detected by quantitative reverse transcription-PCR (qRT-PCR) and western blotting, respectively. The viability of the infected hepatocytes was measured by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. We found that nearly 97% of cells were positive for the hepatocyte marker, CK18. After infection of Ad-PLCγ2 and Ad-PLCγ2 siRNA, more than 99% of hepatocytes expressed GFP significantly, and mRNA and protein expression of PLCγ2 was up-regulated significantly in Ad-PLCγ2 infected hepatocytes, but down-regulated in Ad-PLCγ2 siRNA2 infected cells. The cell proliferation rate decreased in PLCγ2-overexpressing cells, while the rate increased in PLCγ2-silencing cells. We verified that recombinant Ad-PLCγ2 and Ad-PLCγ2 siRNA2 were constructed successfully. These two recombinant vectors promoted or decreased the expression of PLCγ2 in rat hepatocytes and affected the cell proliferation rate, which provides a useful tool for further investigation of the role of PLCγ2 in hepatocyte apoptosis.

Erratum to: Study on injury effect of food additive citric acid on liver tissue in mice

Unfortunately, in the original publication of the article, the citation and the corresponding reference appeared incorrectly as below: Citation: Koca et al. (2005) reported that citric acid could significantly increase micronucleus frequency in red blood cells of Tinca tinca. Reference: Koca YB, Koca S, Yildiz S, Gürcü B, Osanç E, Tunçbaş O, Aksoy G (2005) Investigation of histopathological and cytogenetic effects on Lepomis gibbosus (Pisces: Perciformes) in the Cine stream (Aydin/Turkey) with determination of water pollution. Environ Toxicol 20:560–571 The correct citation and reference are given below: Citation: Koca et al. (2010) reported that citric acid could significantly increase micronucleus frequency in red blood cells of Tinca tinca. Reference: Kocak Y, Gaffaroglu M, Yuksel E (2010) In vivo micronuclei induction by food additive citric acid in peripheral erythrocytes of the fish Tinca tinca. Fresen Environ Bull 19(8a):1608–1614.

Study of antagonism of citric acid on aluminum-induced toxicity in mice testis cells

To study the qualitative changes in testis tissue after aluminum chloride (AlCl3) administration and to determine whether citric acid (CA) has a protective effect against testis damage induced by AlCl3. In this study, 80 Kunming white mice were randomly separated into eight groups: (1) control, (2) CA (120 mg/kg), (3, 4 and 5) AlCl3 (20, 40 and 60 mg/kg), (6, 7 and 8) AlCl3 (20, 40 and 60 mg/kg) plus CA (120 mg/kg). After animals were killed, all testes were histopathologically examined under light microscopy; T-SOD and GSH-Px activities, H2O2 and MDA contents, and Bax and Bcl-2 levels were detected with the corresponding assay kits; DNA fragmentation were electrophoretically examined. Histopathological results indicated that AlCl3 severely damage to mouse testis tissues, however, the protective effects on testes was observed when AlCl3 combined with CA. Biochemical examination suggested that T-SOD and GSH-Px activities significantly decreased (P<0.05) in AlCl3 groups, while remarkably improved in CA+AlCl3 (especially middle and high dose groups) groups; H2O2 and MDA levels in higher-dose AlCl3 groups were obviously higher (P<0.05) than in CA+ corresponding dose AlCl3 groups. Cell apoptosis assays showed that Bax/Bcl-2 ratio in higher-dose AlCl3 groups were higher than CA+ higher-dose AlCl3 groups; and DNA fragmentation in middle-dose AlCl3 groups was much more apparent in CA+ middle-dose AlCl3 groups. It can be concluded that a certain concentration of AlCl3 exerts a reproductive toxicity to mice, and administration of citric acid can reduce the adverse effects of AlCl3 on testis.