Expression of calmodulin-related genes in lead-exposed mice.

The toxic metal lead is a widespread environmental polutant that can adversely affect human health. However, the underlying mechanisms of lead-induced toxicity are still largely unknown. The mechanism of lead toxicity was presumed to involve cross reaction between Pb(2+) and Ca(2+) with calmodulin dependent systems. The aim of the present study was thus to identify differential expression of calmodulin-related genes in the spleen of lead-exposed mice. We performed microarray analysis to identify differentially expressed genes. RNAs from spleen tissue of lead exposed animals (n=6) and controls (n=6) were converted to labeled cRNA and hybridized to Illumina mouse WG-6_v2_Bead Chip. Expression profiles were analyzed using Illumina BeadStudio Application. Real-time RT-PCR was conducted to validate the microarray data. By microarray analysis 5 calmodulin-related genes (MAP2K6, CAMKK2, CXCR4, PHKA2, MYLK) were found to be differently expressed in lead exposed compared with control mice (p<0.05). The results of Real-time RT-PCR showed that MAP2K6 and CAMKK2 were up-regulated and CXCR4 was down-regulated in lead exposure, but there were no significant differences in PHKA2 and MYLK expression between the lead exposed and control group. These results show that lead exposure produced significant changes in expression of a variety of genes in the spleen and can affect calmodulin-related gene expression.

Lead-induced morphological changes and amyloid precursor protein accumulation in adult rat hippocampus.

Lead is an important environmental pollutant that exerts potent toxic effects on many organs. The toxic effects of lead are less well known for adult brain than for children. We investigated the morphological changes and amyloid precursor protein (APP) accumulation in the adult rat hippocampus following exposure to lead. Forty rats were divided into two groups of 20. One group was exposed to 580 parts per million (ppm) lead acetate and other group to an identical concentration of sodium acetate as a control group. After exposure to lead for 3 months, the hippocampus was examined by electron microscopy and APP levels in the hippocampus were detected using immunohistochemistry. Lead levels in the blood of rats exposed to lead were significantly higher than in the controls. The morphological changes in the hippocampus included mitochondrial degeneration, apoptosis and abnormal synapses in the rats exposed to lead. APP in hippocampus was increased significantly in the group exposed to lead compared to controls. We determined that lead exposure causes accumulation of APP and morphological changes in the adult rat hippocampus.

[Proliferation of the mesenchymal stem cells in a delayed fat flap: an experimental study in rabbits].

OBJECTIVE: To investigate the possibility to enhance the proliferation of adipose-derived stem cells (ASCs) in a delayed fat flap in rabbits. METHODS: A delayed fat flap was formed in one side of inguinal region of a rabbit. 21 days after operation, the fat tissues at the delayed flaps and at the unoperated side were harvested and digested with 0.25% collagenase and sieved. The cell suspensions were centrifuged. The cells were obtained from tissue precipitate after centrifugation. The expression rates of the surface marker (CD29, CD44, CD14 and CD45) were measured by FCM and compared between the experimental and control groups. RESULTS: Expression rates of CD29 and CD44 were higher in the delayed fat flap (74.06% and 90.74%) than in the contralateral fat tissue (62.88% and 77.54%, P < 0.05), while those of CD14 and CD45 were lower in the delayed fat flap (57.66% and 4.84%) than in the contralateral fat tissue (72.10% and 75.82%, P < 0.05 and P < 0.01). CONCLUSIONS: Tissue hypoxic ischemia such as fat tissue in a delayed fat flap can promote proliferation of ASCs. It indicates that tissue in the delayed flap may be transplanted with better survival rate. The ischemia pretreatment of fat tissue may become a new method for fat transplantation.

Analysis of the nuclear localization signal of TRF1 in non-small cell lung cancer.

Several studies revealed a similar down-regulation of telomeric repeat binding factor 1 (TRF1) in tumors. We have previously reported the TRFl expression levels were down-regulation in non-small cell lung cancer (NSCLC). The regulation of TRFl localization is proposed to be important for the function and expression. The nuclear localization signal (NLS) and nuclear export signal (NES) are often important clues to localization of protein. The objective of the present study was to investigate the NLS and NES of TRFl in NSCLC patients. Thirty (30) patients with NSCLCs had undergone radical operations in The First Affiliated Hospital, College of Medicine, Zhejiang University. DNA sequences of NLSs and NESs were amplified by PCR. The PCR products were analyzed by DNA sequencing. There were four NLSs of the TRFl protein, including two monopartite and two bipartite NLSs. The NLSs sequences were included in 337KKERRVGTPQSTKKKKESRR356. The exon 8 and exon 9 of TRFl DNA were covered the NLS sequences. The sequences of predicted NESs were 11WMLDFLCLSL86 and 174NLLKLQALAV183, respectively. The exon 1, exon 3 and exon 4 of TRFl were covered the NES sequences. In NSCLCs, there was no a mutation, deletion, or substitution in NLS and NES of TRFl. We conclude that the NLS and NES sequences in NSCLCs patients did not have mutations. Down-expression of TRFl does not indicate gene mutation of NLS and NES in NSCLCs.

[Characteristics of genesis and development of peritoneal adhesion by different causes: experiment with rats].

OBJECTIVE: To investigate the characteristics of genesis and development of peritoneal adhesion by different causes. METHODS: 236 rats underwent laparotomy with their vermiform processes lifted up and were randomly divided into 5 groups: Group A (control group), with the vermiform process exposed to air for 5 min, Group B, with the vermiform process smeared with talcum powder; Group C, with the vermiform process scraped by scalpel; Group D, with the tip of vermiform process stabbed by needle so as to squeeze the contents of intestine to cause infection; and Group E, with the artery of vermiform process ligated. Then the abdominal incision was sutured. 1, 2, 4, and 6 weeks after the treatment 11-12 rats from each group were randomly to undergo laparotomy. The degree of adhesion was graded blindly by Bhatia's method. The vermiform process was resected to undergo pathological examination and examination of the level of organ hydroxyproline (OHP) was detected. RESULTS: (1) At different time points the adhesive grades of Groups B-E were all significantly higher than that of Group A (all P < 0.05) and the adhesive grades of Groups B and D were both significantly higher than those of Groups C and E (both P < 0.05). There were no significant differences in the adhesive degree 1, 2, and 4 weeks after the treatment between Groups C and E, however, the adhesive degree of Group E was significantly lower than that of Group C (P < 0.05). (2) There were not significant differences in the OHP levels at any time points in Group A (all P > 0.05). There were not significant differences in the OHP levels 1, 2, and 4 weeks after the treatment (all P > 0.05), and the levels 8 weeks after the treatment were all significantly lower than those 1, 2, and 4 weeks after the treatment (all P < 0.05) in Groups B, D, and E. In Group C the OHP level 2 weeks after the treatment was 0.275 +/- 0.031 microg/mg protein, significantly lower than that 1 week after (0.221 +/- 0.036 microg/mg protein, P < 0.05), and the OHP level 8 weeks after the treatment was 0.254 +/- 0.039 microg/mg protein, significantly lower than those 1, 2, and 4 weeks after (all P < 0.05). The OH levels 1, 2, and 4 weeks after the treatment of the 4 experimental groups were all significantly higher than that of the control group (all P < 0.05). 8 weeks after the treatment the level of OHP of Groups B was significantly higher than that of Group A (P < 0.05), however, the OHP levels of Group C, D, and E had all decreased to almost similar to that of Group A (all P > 0.05). (3) The adhesive degrees of Groups C and E were significantly positively correlated with the OHP level (both P < 0.05), however, the adhesive degrees of Groups B and D were not significantly correlated with the OHP level (both P > 0.05). The adhesive degrees 1, 2, and 4 weeks after the treatment of the 5 groups were all significantly positively correlated with the OHP levels (all P < 0.05, however, the adhesive degrees 8 weeks after the treatment of the 5 groups were all not significantly correlated with the OHP levels (all P > 0.05). (4) The main pathological changes of Group B were foreign body granuloma reaction and fibroplasia in Group B and unspecific inflammatory reaction and fibroplasia in Groups C, D, and E. CONCLUSION: Abdominal adhesions resulting from different causes show different characteristics. The abdominal adhesion caused by foreign bodies and that caused by infection are relatively severe and more difficult to recover than those caused by injury and ischemia. It is more reliable to use OHP level as a marker of abdominal adhesion in the early stage.

Therapeutic effect of mild hypothermia on severe traumatic head injury.

OBJECTIVE: To investigate the therapeutic effect of mild hypothermia on severe traumatic brain injury. METHODS: Eighty-six in-patients with severe traumatic brain injury treated ordinarily were consecutively randomized into two groups: a hypothermia group (n=43) and a normothermia group (the control group, n=43). In the hypothermia group, the core temperature (i.e., nasopharyngeal or brain temperature) of the patient was reduced to and maintained at 33-35 degrees C with a systemic cooling blanket. Natural rewarming began after 3-5 days (mean: 4.3 days) of hypothermia treatment. In the control group, the patient received no hypothermia treatment. The vital sign, extradural pressure and serum superoxide dismutase were observed and measured during treatment, and the complications as well as the Glasgow outcome scale were evaluated at 2 years after injury. RESULTS: The mean extradural pressure in the hypothermia group (27.38 mm Hg +/- 4.88 mm Hg at 24 hours, 29.40 mm Hg +/- 4.50 mm Hg at 48 hours and 26.40 mm Hg +/- 4.13 mm Hg at 72 hours after injury) was much lower than that in the control group (32.63 mm Hg +/- 3.00 mm Hg, 34.80 mm Hg +/- 6.00 mm Hg and 31.81 mm Hg +/- 4.50 mm Hg respectively at 24, 48 and 72 hours, P<0.05). The mean serum superoxide dismutase levels in the hypothermia group on days 3 and 7 (583.7 microg/L +/- 99.6 microg/L and 699.4 microg/L +/- 217.3 microg/L, respectively) were much higher than those in the control group at the same time period (446.6 microg/L +/- 79.5 microg/L and 497.1 microg/L +/- 101.2 microg/L, respectively, P<0.01). The recovery rates at 2 years after injury were 65.1% in the hypothermia group and 37.2% in the control group (P<0.05). The mortality rates were 25.6% in the hypothermia group and 51.2% in the control group (P<0.05). The complications, including pulmonary infections, thrombocytopenia (platelet count < 100 x 10(9)/L), hemorrhage in the digestive tract, electrolyte disorders and renal malfunction, were managed without severe sequelae. CONCLUSIONS: Mild hypothermia is a safe and effective therapeutic method, which can lower the extradural pressure, increase the serum superoxide dismutase and improve the neurological outcomes without severe complications in the patients with severe traumatic brain injury.