Identification of G2/M phase transition by sequential nuclear and cytoplasmic changes and molecular markers in mice intestinal epithelial cells.

Although the regulatory network of G2/M phase transition has been intensively studied in mammalian cell lines, the identification of morphological and molecular markers to identify G2/M phase transition in vivo remains elusive. In this study, we found no obvious morphological changes between the S phase and G2 phase in mice intestinal epithelial cells. The G2 phase could be identified by Brdu incorporation resistance, marginal and scattered foci of histone H3 phosphorylated at Ser10 (pHH3), and relatively intact Golgi ribbon. Prophase starts with nuclear transformation in situ, which was identified by a series of prophase markers including nuclear translocation of cyclinB1, fragmentation of the Golgi complex, and a significant increase in pHH3. The nucleus started to move upwards in the late prophase and finally rounded up at the apical surface. Then, metaphase was initiated as the level of pHH3 peaked. During anaphase and telophase, pHH3 sharply decreased, while Ki67 was obviously bound to chromosomes, and PCNA was distributed throughout the whole cell. Based on the aforementioned markers and Brdu pulse labeling, it was estimated to take about one hour for most crypt cells to go through the G2 phase and about two hours to go through the G2-M phase. It took much longer for crypt base columnar (CBC) stem cells to undergo G2-prophase than rapid transit amplifying cells. In summary, a series of sequentially presenting markers could be used to indicate the progress of G2/M events in intestinal epithelial cells and other epithelial systems in vivo.

Regulation of Embryonic Stem Cell Self-Renewal and Differentiation by MicroRNAs.

MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression. They play an important role in various cellular processes such as apoptosis, differentiation, secretion, and proliferation. Embryonic stem cells (ESCs) are derived from the inner cell mass of the blastocyst stage of the embryo. miRNAs are critical factors for the self-renewal and differentiation of ESCs. In this review, we will focus on the role of miRNAs in the self-renewal and directional differentiation of ESCs. We will present the current knowledge on key points related to miRNA biogenesis and their function in ESCs.

P68 RNA helicase as a molecular target for cancer therapy.

The DEAD-box family of RNA helicase is known to be required in virtually all cellular processes involving RNA, and p68 is a prototypic one of the family. Reports have indicated that in addition to ATPase and RNA helicase ability, p68 can also function as a co-activator for transcription factors such as estrogen receptor alpha, tumor suppressor p53 and beta-catenin. More than that, post-translational modification of p68 including phosphorylation, acetylation, sumoylation, and ubiquitylation can regulate the coactivation effect. Furthermore, aberrant expression of p68 in cancers highlights that p68 plays an important role for tumorgenesis and development. In this review, we briefly introduce the function and modulation of p68 in cancer cells, and put forward envisagement about future study about p68.

Detection of CD4+ CD25+ FOXP3+ regulatory T cells in peripheral blood of patients with chronic autoimmune urticaria.

BACKGROUND/OBJECTIVES: Compelling evidence indicates a significant role for a population of CD4(+) T regulatory cells in suppressing immune responses and in maintaining immunological homeostasis. This study aims to investigate the potential role of CD4(+) CD25(HIGH) FOXP3(+) T regulatory cells in patients with chronic autoimmune urticaria and to define the characteristics of CD4(+) CD25(HIGH) FOXP3(+) cells in chronic urticaria. METHODS: We used flow cytometry to assess the expression of CD4(+) CD25(HIGH) FOXP3(+) cells in the peripheral blood mononuclear cells of patients with chronic autoimmune urticaria. RESULTS: In this study, we found that patients with chronic autoimmune urticaria have a significantly reduced frequency of CD4(+) CD25(HIGH) FOXP3(+) cells (1.39 ± 0.27% vs 2.09 ± 0.34%; P = 0.001) in their peripheral blood, accompanied by a decreased intensity of FOXP3 expression (50.13 ± 9.79 vs 68.19 ± 6.40; P < 0.001). Notably, although patients with chronic idiopathic urticaria had a reduced frequency of CD4(+) CD25(HIGH) FOXP3(+) cells (1.85 ± 0.46% vs 3.64 ± 0.48%; P < 0.001), their FOXP3 expression levels did not differ from those in healthy controls. CONCLUSIONS: Patients with chronic autoimmune urticaria displayed a reduced percentage of CD4(+) CD25(+) FOXP3(+) regulatory T cells. The results imply CD4(+) CD25(+) FOXP3(+) regulatory T cells may contribute to the autoimmune pathological process of chronic autoimmune urticaria.

Experimental research on the management of combined radiation-burn injury in China.

Combined radiation-burn injury can occur in people exposed to nuclear explosions, nuclear accidents or radiological terrorist attacks. Using different combined radiation-burn injury animal models, the pathological mechanisms underlying combined radiation-burn injury and effective medical countermeasures have been explored for several years in China, mainly at our institute. Targeting key features of combined radiation-burn injury, several countermeasures have been developed. Fluid transfusion and the calcium antagonist verapamil can prevent early shock and improve myocardial function after combined radiation-burn injury. Recombinant human interleukin 4 (rhIL-4) is able to effectively reduce bacterial infection and increase intestinal immunological ability. Chitosan-wrapped human defensin 5 (HD5) and glucagon-like peptide 2 (GLP-2) nanoparticles can increase the average survival time of animals with severe combined radiation-burn injury. After treatment by cervical sympathetic ganglia block (SB), hematopoietic function is promoted and the release of inflammatory cytokines is suppressed. The optimal time for escharectomy and allo-skin grafting is 24 h after injury. Transfusion of irradiated (20 Gy) or stored (4°C, 7 days) blood improves the survival of allo-skin grafting and allo-bone marrow cells. In conclusion, as our understanding of the mechanisms of combined radiation-burn injury has progressed, new countermeasures have been developed for its treatment. Because of the complexity of its pathology and the difficulty in clinical management, further efforts are needed to improve the treatment of this kind of injury.

Current state and future perspectives of trauma care system in mainland China.

OBJECTIVE: To investigate the current state of trauma care in mainland China, and to propose possible future suggestions for the development of the trauma care system in mainland China. METHOD: An extensive Medline/PubMed search on the topic of trauma care or trauma care system was conducted. Publications in Chinese that could best describe the state of trauma care in China were also included. In addition, two meetings were held by Group for Trauma Emergency Care and Multiple Injuries, Trauma Society of Chinese Medical Association to discuss the development and perspectives of trauma care system in mainland China. Important conclusions from the two meetings were included in this publication. RESULTS: Trauma has become an increasing public health problem in mainland China in association with the rapid growth of the economy over the past 30 years. Although great progress has been made in regards to the care of the injured, there is still no government agency dedicated to deal with trauma-related issues, or a national trauma care system operating on the Chinese mainland. Various trauma prevention measures have been taken, but with little effect. Funds contributed to trauma-related research has increased in recent years and promoted rapid development in this field, but further improvement in research is needed. However, many groups such as the Trauma Society of the Chinese Medical Association have continued to explore mechanisms for the treatment of trauma patients and have developed various types of regional trauma care systems, resulting in improved trauma care and a better outcome for the injured. CONCLUSIONS: Although great progress has been made in trauma care in mainland China, there are many failings. To improve trauma care in China, the establishment of a sophisticated trauma system and various enhancements on trauma prevention are urgently required.

Protective effect of atorvastatin on radiation-induced vascular endothelial cell injury in vitro.

Vascular endothelial cells are very sensitive to ionizing radiation, and it is important to develop effective prevent agents and measures in radiation exposure protection. In the present study, the protective effects of atorvastatin on irradiated human umbilical vein endothelial cells (HUVEC) and the possible mechanisms were explored. Cultured HUVEC were treated by atorvastatin at a final concentration of 10 µ mol/ml for 10 minutes, and then irradiated at a dose of 2 Gy or 25 Gy. Twenty-four hours after irradiation, apoptosis of HUVEC was monitored by flow cytometry, and the expression of thrombomodulin (TM) and protein C activation in HUVEC was respectively assessed by flow cytometry and spectrophotometry. After treatment with atorvastatin for 24 h, the rate of cell apoptosis decreased by 6% and 16% in cells irradiated with 2 Gy and 25 Gy, respectively. TM expression increased by 77%, 59%, and 61% in untreated cells, 2 Gy irradiation-treated cells, and 25 Gy irradiation-treated cells, respectively. The protein C levels in 2 Gy and 25 Gy irradiation-treated cells were reduced by 23% and 34% when compared with untreated cells, but up-regulated by 79% and 76% when compared with cells which were irradiated and treated with atorvastatin. In conclusion, these data indicate that atorvastatin exerts protective effects on irradiated HUVEC by reducing apoptosis by up-regulating TM expression and enhancing protein C activation in irradiated HUVEC.

Protective effect of an extract from Periplaneta americana on hematopoiesis in irradiated rats.

PURPOSE: To investigate the protective effect of W(11)-a(12), an extract from Periplaneta americana, on hematopoiesis in irradiated rats. MATERIALS AND METHODS: Wistar rats receiving total body irradiation of (60)Co gamma-rays alone or with combined radiation and skin wound injury were used in this study. W(11)-a(12) was applied either topically into the skin wounds or systemically by intraperitoneal injection. The numbers of white blood cells in peripheral blood, the nucleated cells and the colony-forming unit of granulocyte/macrophage progenitors (CFU-GM) in bone marrow were measured, respectively. RESULTS: Topical application of W(11)-a(12) into skin wounds in rats with combined 6 Gy total body irradiation and skin wound injury could increase the neutrophils and macrophages in the wounded area and the nucleated cells in bone marrow at 24 h and 48 h, while the peripheral white blood cells did not show significant change. However, in rats with 4 Gy total body irradiation alone, the peripheral white blood cells, bone marrow nucleated cells and the number of colony-forming unit of granulocyte-macrophage progenitors were all significantly higher in the treatment groups by intraperitoneal injection of W(11)-a(12) than those in the control groups by injection of normal saline at days 3 and days 5 after radiation. CONCLUSIONS: W(11)-a(12) showed a protective effect on hematopoiesis after total body irradiation and could increase the inflammatory cells in wounded tissues at the initiation stage after irradiation, which will benefit the management of combined radiation and skin wound injury.

CXCR4 gene transfer enhances the distribution of dermal multipotent stem cells to bone marrow in sublethally irradiated rats.

Our previous study indicated that systemically transplanted dermal multipotent cells (DMCs) were recruited more frequently to bone morrow (BM) of rats with sublethal irradiation than that of normal rats, and the interactions between stromal-derived factor (SDF-1) and its receptor (CXC chemokine receptor 4, CXCR4) played an important role in this process. In the present study, we aimed to investigate whether CXCR4 gene transfer could promote the distribution of DMCs into irradiated BM and accelerate its function recovery. Firstly, adenovirus vector of CXCR4 (Adv-CXCR4) and green fluorescent protein (Adv-GFP) were constructed. Then male DMCs infected by Adv-CXCR4 (group A), or infected by Adv-GFP (group B), and non-infected DMCs (group C) were transplanted into irradiated female rats, and real-time polymerase chain reaction for the sex-determining region of Y chromosome was employed to determined the amount of DMCs in BM. The functional recovery of BM was examined by hematopoietic progenitor colonies assay. The results showed that the amount of DMCs in BM of group A was greater than that in group B and group C from day 5 after injury (P < 0.05), and the amount of CFU-F, CFU-E and CFU-GM were greater than that in group B and group C from day 14 after injury (P < 0.05). These findings suggest that DMCs infected by Adv-CXCR4 distributed more frequently to the bone marrow of sublethally irradiated rats and could accelerate hematopoiesis function recovery.

[Studies on the treatment of combined radiation-burn injury].

Combined radiation-burn injuries mainly occur under the circumstances of nuclear explosion, nuclear accident, nuclear terrorism, depleted uranium attack, as well as secondary injuries following attack on nuclear installation. Combination of burn and radiation injuries bring along more serious whole body damage, more complicated pathological mechanism and much more difficult management. Research progress on the pathological mechanism and medical management of several key links of combined injury were discussed in this paper. (1) Enhancement of early first aid and prevention of early death of wounded. (2) Damage and restoration of hemopoietic function. (3) Disturbance of immune function and prevention and treatment of infection (mainly on the intestinal mucosa immunity and enterological infection). (4) Management of burn wound. (5) The role of several important measures in the comprehensive treatment.

Recruitment of transplanted dermal multipotent stem cells to sites of injury in rats with combined radiation and wound injury by interaction of SDF-1 and CXCR4.

Systemic transplantation of dermal multipotent stem cells has been shown to accelerate both hematopoietic recovery and wound healing in rats with combined radiation and wound injury. In the present study, we explored the mechanisms governing the recruitment of dermal multipotent stem cells to the sites of injury in rats with combined injury. Male dermal multipotent stem cells were transplanted into female rats, and using quantitative real-time PCR for the sex-determining region of Y chromosome, it was found that the amounts of dermal multipotent stem cells in irradiated bone marrow and wounded skin were far greater than those in normal bone marrow and skin (P < 0.01). However, incubation of dermal multipotent stem cells with AMD3100 before transplantation, which specifically blocks binding of stromal cell-derived factor 1 (SDF-1) to its receptor CXCR4, diminished the recruitment of dermal multipotent stem cells to the irradiated bone marrow and wounded skin by 58 +/- 4% and 60 +/- 4%, respectively (P < 0.05). In addition, it was confirmed that the expression of SDF-1 in irradiated bone marrow and wounded skin was up-regulated compared to that in their normal counterparts, and in vitro analysis revealed that irradiated bone marrow and wounded skin extracts had a strong chemotactic effect on dermal multipotent stem cells but that the effect decreased significantly when dermal multipotent stem cells were preincubated with AMD3100 (P < 0.05). These data suggest that transplanted dermal multipotent stem cells were recruited more frequently to the irradiated bone marrow and wounded skin than normal bone marrow and skin and that the interactions of SDF-1 and CXCR4 played a crucial role in this process.

Autoantibodies to the high-affinity IgE receptor in patients with asthma.

Autoimmune diseases have been implicated as a cause of intrinsic asthma; however, there is little data on the role of autoimmunity in the pathogenesis of asthma. The purpose of this study was to investigate circulating autoantibodies against the high-affinity IgE receptor Fc(epsilon)RI in patients with asthma. Seventy-eight patients with asthma and 32 healthy individuals as control subjects were included. All subjects were tested with basophil histamine releasing assay and immunoblotting to assess for the potential presence of receptor Fc(epsilon)RI autoantibodies. Of the 78 asthma patients total subjects, 25 (32.1%) had a positive by basophil histamine releasing assay and 23 (29.5%) by immunoblotting. Both of them were significant higher than the positive rate, 9.4% (p < 0.05) and 9.4% (p < 0.05), respectively. Our data demonstrated that aberrant autoantibodies against the high-affinity IgE receptor Fc(epsilon)RI were found in some patients with asthma implies that the autoimmunity may be one factor in intrinsic asthma pathogenesis.

Effects of peritoneal lavage fluid from radiation or/and burn injured rats on the growth of hematopoietic progenitor cells.

PURPOSE: To evaluate the effects of peritoneal lavage fluids from radiation injury, burn injury and combined radiation-burn injury on the growth of hematopoietic progenitor cells (HPC). MATERIALS AND METHODS: Rats were divided into four groups: A radiation group (RG), a burn group (BG), a combined radiation-burn group (CRBG) and normal control group (NG). RG and CRBG rats were irradiated with 12 Gy, and burns of 30% total body surface area were generated in group BG and group CRBG. Peritoneal lavage fluids were collected and tested for their effects on the growth of erythrocyte progenitor cells or granulocyte-macrophage progenitor cells of BALB/c mice in vitro. RESULTS: The numbers of colony forming units-erythroid (CFU-E), burst forming units-erythroid (BFU-E) and colony-forming units-granulocyte-macrophage (CFU-GM) formed after treatment with lavage fluids from BG or CRBG were significantly higher than those from NG. However, fewer CFU-E, BFU-E or CFU-GM colonies were found after treatment with lavage fluid from the RG. In lavage fluid from BG and CRBG, the concentration of interleukin-6 (IL-6), interleukin-8 (IL-8) and tumor necrosis factor alpha (TNFalpha) was increased in comparison to NG and RG. Treatment with these cytokines had similar promoting effects on the growth of hematopoietic colonies and neutralizing antibodies inhibited these effects significantly. CONCLUSIONS: Burns increase the responsiveness of the system and help the proliferation of hematipoietic progenitor cells, while radiation decreases all these responses relative to both the controls and the burn plus radiation group.

Effects of serum from rats with combined radiation-burn injury on the growth of hematopoietic progenitor cells.

BACKGROUND: This study aims to observe the effects of blood serum from rats with radiation injury, burn injury, and combined radiation-burn injury on the growth of hematopoietic progenitor cells and to explore the possible mechanisms. METHODS: Serum from rats with radiation injury, burn injury, and combined radiation-burn injury were collected at 3 hours, 12 hours, 24 hours, 48 hours, 72 hours, and 96 hours after injury and then was added to the culture medium to see its effect on the growth of hematopoietic progenitor cells (HPCs) at a final protein concentration of 10 microg/mL. Radioimmunoassay and enzyme-linked immunosorbent assay were employed to measure the level of tumor necrosis factor (TNF)-alpha and interleukin (IL)-6 in each group, and the effect of TNF-alpha and IL-6 on the growth of HPC was also observed. RESULTS: The number of HPCs colonies formed after addition of the serum from rats with burn or combined radiation-burn injuries was significantly higher than that from normal rats at 3 hours, 12 hours, 24 hours, 48 hours, 72 hours, and 96 hours after injury and reached its peak value at 24 hours after injury. However, fewer HPCs colonies were found after the addition of the serum from irradiated rats. At the same time, the levels of TNF-alpha and IL-6 in the serum of burn group and combined radiation-burn injury group were significantly higher than that of normal group, and much higher than that of the irradiation injury group (p < 0.01). Also, TNF-alpha and IL-6 demonstrated promoting effect on the growth of HPC. CONCLUSION: Serum from rats with burn injury and combined radiation-burn injury stimulates the growth of HPCs, while serum from irradiated rats shows inhibitory effects on the growth of HPCs. These effects may lie in the different level of TNF-alpha and IL-6 in the serum of each group.

Crucial role of SDF-1/CXCR4 interaction in the recruitment of transplanted dermal multipotent cells to sublethally irradiated bone marrow.

Our previous study indicated that dermal multipotent cells (DMCs) could engraft into bone morrow (BM) of rats with sublethal irradiation and promote hematopoietic recovery after being transplanted systemically, but the mechanisms determining the recruitment of DMCs to the irradiation injured BM remain unclear. In the present study, we investigated the role of stromal cellderived factor-1 (SDF-1)/CXCR4 interaction in this process. Male DMCs were isolated and transplanted into female rats systemically, and by employing quantitative real-time TaqMan polymerase chain reaction for the sex-determining region of Y chromosome, it was found that the amount of DMCs in BM of rats with sublethal irradiation was about 3 times more than that of normal rats (P < 0.01). Incubation of DMCs with AMD3100 before transplantation, which specifically blocks binding of SDF-1 to its endogenous receptor CXCR4, diminished recruitment of DMCs to the injured BM by 57.2 +/- 5.5% (P < 0.05). In addition, it was confirmed that the expression of SDF-1 in injured BM was up-regulated when compared with that in normal BM, and in vitro analysis revealed that BM extracts from irradiated rats had a strong chemotactic effect on DMCs, which decreased significantly when DMCs were pre-incubated with AMD3100 (P < 0.05). These data suggest that transplanted DMCs were recruited more frequently to irradiation-injured BM than normal BM and the interactions of SDF-1/CXCR4 played an important role in this process.

[Effects of blood serum from rats with combined radiation-thermal injury on the bone marrow hematopoietic progenitor cells growth].

To observe the effects of blood serum from rats with radiation injury, thermal injury and combined radiation-thermal lesions on growth of hematopoietic progenitor cells and the change of their serum cytokine levels, total body irradiation of rats was performed with 12 Gy gamma ray from a (60)Co source, and 30% total body surface area III degree thermal lesion on the back was inflicted with a 5 kW bromotungsten lamp. The blood serum from these animals was collected at 3, 12, 24, 48, 72 and 96 hours after injury. Then the blood serum was added to the culture medium of erythrocyte progenitor cells (CFU-E, BFU-E) or granulocyte-macrophage progenitor cells (CFU-GM) at final concentration of 10 microg/ml. The results showed that the colony number of CFU-E, BFU-E and CFU-GM formed after addition of the blood serum from rats with thermal or combined radiation-thermal injury was significantly higher than that from normal rats at 3, 12, 24, 48, 72 and 96 hours after injury and reached its peak value at 24 hours after injury (342.8, 261.6 and 228.4% respectively from burned rats, 252.4, 205.1 and 174.2% respectively from rats with combined radiation-thermal injury as compared with that of normal rats). However, a few CFU-E, BFU-E or CFU-GM formation was found after addition of the blood serum from irradiated rats. At the same time, the level of TNF alpha and IL-6 in serum of burn group and combined radiation-thermal injury group was markedly higher than that of normal group, even more higher than that of irradiation injury group (P < 0.01). It is concluded that the blood serum from rats with thermal lesion or combined radiation-thermal injury improves the growth of erythrocyte and granulocyte progenitor cells. On the contrary, the blood serum from the irradiated rats shows the inhibiting effects, definitely related to their serum cytokines changes.

[Effects of exogenerous nucleotides on the apoptosis of intestinal epithelial cells IEC-6].

OBJECTIVE: To investigate the effects of exogenous nucleotides on apoptosis of a normal rat small intestinal epithelial cell line, IEC-6. METHODS: Cultured IEC-6 cells were treated by four kinds of monophosphate nucleotides and their mixture prepared according to their composition in human milk, then the cell apoptosis was determined by flow cytometry measurement, morphologic characterization, and electron-microscope observation. RESULTS: IEC-6 cells treated with AMP or GMP showed a apotosis peak in flow cytometry measurement, but only AMP produce typical apoptosis characteristics in electron-microscope observation. Pyrimidine nucleotides (UMP and CMP)and nucleotides mixture could not induce apoptosis. However, UMP could significantly eliminate the apoptosis-inducing effects of AMP or GMP. CONCLUSION: Purine nucleotides induce apoptosis of IEC-6, inducing effects of purine nucleotides. pyrimidine nucleotides UMP could abolish the apoptosis-inducing effects of purine nucleotides.

[Effect of lipopolysacharide on the biological features and growth factor secretion power of U937 cell line].

OBJECTIVE: To investigate the effect of lipopolysacharide (LPS) in different concentrations on the biological features and growth factor secretion power of U937 cell line. METHODS: In vitro cultured U937 cells were stimulated by 0 (as control), 0.1, 1.0, 10.0, 50.0 and 100 micro g/ml LPS respectively for 24 hours. Thereafter, the cell proliferation ability was determined by MTT method. The cell apoptosis rate was determined by flow cytometry. The changes in the contents of transforming growth factor beta(1) (TGFbeta(1)) and vascular endothelial growth factor (VEGF) of the supernatant of the cell culture were assessed by ELISA. RESULTS: Apoptosis and TGFbeta(1) secretion could be induced by LPS in dose of 0.1 to 100 micro g/ml when compared with that without LPS challenge (P < 0.05 - 0.01). In detail, LPS in lower dose (0.1, 1.0 and 10.0 micro g/ml) could promote the proliferation of U937 (P < 0.05 - 0.01) but exerted no effect on VEGF secretion. In contrary, LPS in high dose (50 and 100 micro g/ml) could promote VEGF secretion (P < 0.01) but exerted no effects on the proliferation of U937 cells. CONCLUSION: U937 cells could be activated to increase the secretion of TGFbeta(1) by LPS in optimal dose of 0.1 - 10.0 micro g/ml, but the secretion of VEGF could only be promoted by LPS in higher concentration.

Transplantation of dermal multipotent cells promotes the hematopoietic recovery in sublethally irradiated rats.

Our previous study indicated that dermal multipotent cells with the differentiation capacity to form cells with the phenotypic properties of osteocytes, adipocytes, chondrocytes, and neurons in specific inducing media could be isolated from the enzymatically dissociated dermal cells of newborn rats by their adherence to culture dish plastic. We have also observed that the systemic transplantation of dermal multipotent cells could not repopulate the hematopoietic system in lethally irradiated rats. In this paper, we found that a transplantation of plastic-adherent dermal multipotent cells into sublethally irradiated rats led to a significant increase of white blood cells in peripheral blood, nucleated cells, CFU-GM, and CFU-F colonies in bone marrow. FISH analysis, using a Y-chromosome specific probe, showed that dermal multipotent cells could engraft into bone marrow in recipients. Flow cytometry (FACS) analysis also showed that the proportion of CD2 and CD25 positive lymphocytes in peripheral blood did not change significantly in two weeks after transplantation. By these results, we infer that dermal multipotent cells may represent an alternative origin of mesenchymal stem cells to restore marrow microenvironment and promote the survival, engraftment, and proliferation of hematopoietic cells.

[The role of homeobox B2 gene in vascular endothelial proliferation and the protective effects of VEGF on the endothelia against radiation injury].

OBJECTIVE: To explore the role of HOXB2 gene in the proliferation of primary human umbilical vein endothelial cells (HUVECs) and the protective effects of VEGF on the endothelia against radiation injury. METHODS: HUVECs were isolated, cultured, subcultured and identified. (1) Liposome coated oligodeoxynucleotide (odn) and homeoboxB2 antisense oligodeoxyncleotide (HOXB2asodn) were prepared prepared in the concentrations of 0.25, 0.5, 1.0 and 2.5 mg/L for the stimulation of HUVEC. (3)H-TdR incorporation test and MTT method were employed to determine the proliferation activity of HUVECs after activation. The cell cycle analysis of HUVECs was determined by flow cytometry. The expression level of HOXB2mRNA within HUVECs was detected by RT-PCR (reverse transcription polymerase chain reaction). (2) HUVECs were separately treated with the addition of VEGF in concentration of 50 microg/L, by radiation in the dose of 6 Gy or 12 Gy (60)Co gamma gamma ray, or radiation with 12 Gy (60)Co gamma gamma ray followed by the addition of VEGF in dose of 50 microg/L. The cellular morphology was observed and the cellular proliferation activity was determined by MTT method. RESULTS: (1) The proliferation activity of HUVECs could be markedly inhibited by liposome coated HOXB2asodn in comparison to liposome-odn (P < 0.05 or 0.001), and the inhibition effect was positively correlated with the increase in asodn concentration. The cell ratio in S phase and the expression level of the HOXB2mRNA could be lowered by asodn in dose of 2.5 mg/L (P < 0.05 or 0.001). (2) Radiation by (60)Co gamma ray could lead to the nuclear enlargement, vacuolation in the cytoplasm, multiplicity of nucleus and nuclear swelling. The proliferative activity of HUVECs was increased from 0.365 +/- 0.047 and 0.487 +/- 0.022 without radiation to 0.557 +/- 0.042 and 0.648 +/- 0.021 24 and 48 hours after 6 Gy radiation However it was decreased to 0.263 +/- 0.038 and 0.306 +/- 0.024 (P < 0.01) after 12 Gy (60)Co gamma ray radiation. Nevertheless, the cell morphology was obviously improved and the proliferation was enhanced by the addition of VEGF after 12 Gy radiation. CONCLUSION: HOXB2 gene played important roles in the biological activities of HUVECs. Small dose (6 Gy) gamma-radiation could promote, but large dose (12 Gy) could decrease the mRNA expression of HOXB2 gene in HUVECs. In addition, VEGF could protect HUVECs against radiation injury.