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AIM: To investigate the genetic relationship between Hirschsprung’s disease (HD) and intestinal neuronal dysplasia (IND) in Chinese population. METHODS: Peripheral blood samples were obtained from 30 HD patients, 20 IND patients, 18 HD/IND combined patients and 20 normal individuals as control. Genomic DNA was extracted according to standard procedure. Exons 11,13,15,17 of RET proto-oncogene were amplified by polymerase chain reaction (PCR). The mutations of RET proto-oncogene were analyzed by single strand conformational polymorphism (SSCP) and sequencing of the positive amplified products was performed. RESULTS: Eight germline sequence variants were detected. In HD patients, 2 missense mutations in exon 11 at nucleotide 15165 G→A (G667S), 2 frameshift mutations in exon 13 at nucleotide 18974 (18974insG), 1 missense mutation in exon 13 at nucleotide 18919 A→G (K756E) and 1 silent mutation in exon 15 at nucleotide 20692 G→A(Q916Q) were detected. In HD/IND combined patients, 1 missense mutation in exon 11 at nucleotide 15165 G→A and 1 silent mutation in exon 13 at nucleotide 18888 T→G (L745L) were detected. No mutation was found in IND patients and controls. CONCLUSION: Mutation of RET proto-oncogene is involved in the etiopathogenesis of HD. The frequency of RET proto-oncogene mutation is quite different between IND and HD in Chinese population. IND is a distinct clinical entity genetically different from HD.

SELDI mass spectrometry was used to investigate protein expression in sera of patients with gastric cancer and gastritis compared to normal volunteers. Differences in peak morphology and intensity were observed in regions of 5910 Da, 5084 Da, 6640 Da and 8691 Da. Patients with gastric cancer exhibited an up-regulation of the 5910 Da peak and a down-regulation of the 8691 Da peak compared to the healthy volunteers; there was also some bi-partitioning and tri-partitioning at the 5084 Da peak. When comparing the sera of these cancer patients with those of gastritis, the former had an up-regulation of the 5910 Da peak and a down-regulation of the 6640 Da peak. This is the first report showing that SELDI sera analysis may be useful in the screening of gastric lesions.

Human colorectal cancer (lovo) cells were chose to study the anti-tumor effects of SFPS and explore the significance of caspase 3 in the apoptosis of lovo cells induced by SFPS. Inhibition of the cell proliferation was measured by MTT assay. SFPS induced apoptosis of lovo cells was observed by electron microscopy, flow cytometry and DNA electrophoresis. Furthermore, the expressions of caspase 3 mRNA and pro-caspase 3 were tested by RT-PCR and Western blot. SFPS exhibited anti-proliferative activity in the dosage and time-dependent manner. After incubation for 24, 36, 48 and 72 h, the IC50 of SFPS on lovo cells was 375 mg/L, 355 mg/L, 178 mg/L and 60 mg/L, respectively. DNA ladders of lovo cells were showed on agarose gel electrophoresis and the fragments of DNA were integral of 180-200 bp 24 hours after SFPS treatment at the doses of 5 mg/L, 50 mg/L and 300 mg/L. However, mistiness DNA ladder or smear was found in lovo cells treated with 500 mg/L SFPS. When SFPS concentration was 1000 mg/l, DNA ladder disappeared and showed smear entirely. Morphological examination showed chromosomal condensation, karyotheca margination, cell shrinkage and the presence of apoptosis bodies with electron microscopy. With concentration dependent manner, the apoptosis and sub-G1 peaks were observed through flow cytometry, but the cell cycle didn't change obviously. Furthermore, the expression of pro-caspase 3 was decreased and the level of caspase 3 mRNA was increased in the time and dose-dependent manner. It suggests that SFPS may induce the apoptosis of lovo cells in vitro resulting in the inhibition of proliferation. And caspase 3 activation may participate in the processes of the apoptosis of lovo cells induced by SFPS.

To confirm that PCR products with heterozygous mutations contain not only wide-type and mutant homoduplexes, but also two types of heteroduplexes.An insertion-deletion mutation in the exon 1 of KRT9 gene (497delAinsGGCT), which caused Chinese epidermolytic palmoplantar keratoderma (EPPK) was investigated by polymerase chain reaction (PCR), polyacrylamide gel electrophoresis (PAGE) and denaturing high-performance liquid chromatography(DHPLC).Two heteroduplexes and two homoduplexes in the PCR product from the heterozygous mutation of the exon 1 of KRT9 (497delAinsGGCT) were detected.PCR products from KRT9 gene with heterozygous mutations contain two types of heteroduplexes. It is without the need to perform heating and cooling PCR products obtained from heterozygous mutations in advance before the mutation screening steps such as denaturing gradient gel electrophoresis (DGGE), temperature gradient gel electrophoresis (TGGE), conformation-sensitive gel electrophoresis (CSGE), DHPLC and heteroduplex analysis (HA), etc.

Scanning retinal electronmicroscopy studies demonstrated the presence of both twin cones (two identical cones adhered together) and double cones (two non-identical cones adhered together) in the retinae of the major forms of fish, which included actinopterygi, elasmobranch, and teleosts. The complexity of these visual cells in the retinae of different groups of fishes was revealed for the first time and reflected on the diversities of functions of the fish retina in general.

The expressions of VEGF-C mRNA, VEGFR-3 and CD31 were studied in order to investigate the correlation between them and neoangiogenesis, hyperplasia of micro-lymphatics and tumor metastases. 34 cases of prostate cancer tissue and 12 cases of adjacent nontumorous tissue specimens were tested. They were marked by VEGFR-3 and CD31 with immunohistochemistic staining and analyzed with image, the micro-lymphatics count (MLC) and microvessel density (MVD) were counted using Weidner's highest vessel density count method; the expression of VEGF-C mRNA was inspected in situ hybridization. The expression of VEGF-C mRNA was 44.12% positive in 34 cases of prostate cancer, the MLC (8.26 +/- 2.73)mm2 and MVD (74.82 +/- 11.76)mm2 in prostate cancer were significantly higher than those in adjacent nontumorous tissue (MLC, 4.82 +/- 3.48/mm2; MVD, 32.86 +/- 5.41/mm2). In addition, there was a correlation between the expression of VEGF-C mRNA and micro-lymphatics metastases and there was a positive correlation between the expression of VEGFR-3 and CD31. The expressions of VEGF-C mRNA , MLC, MVD in stage III and IV and those who have lymph metastasis were higher than those in stage I and II and those who have no lymph metastasis; the expressions of VEGFR-3 and CD31 in VEGF-C mRNA positive groups were significantly higher than those in negative groups. The difference of histopathologic grading in prostate cancer had no statistical significance. VEGF-C could accelerate the hyperplasia of micro-lymphatics and neoangiogenesis induced by tumor and play an important role in tumor lymph metastases. There was a close correlation between the expressions of VEGFR-3, CD31 and tumor metastases. The increase of MLC and MVD on prostate cancer indicated the hyperplasia of new micro-lymphatics and neoangiogenesis in the tumor tissue, which could also be a signal to determine the tumor metastases in clinic.

The aim of this study was to examine microsatellite instability (MSI) and loss of heterozygosity (LOH) of locus D17S396 on chromosome 17 and their influence on the expression of nm23H1 in the epithelial ovarian tumors, which may provide experimental basis for the mechanism of nm23H1 gene and tumor metastasis. Techniques such as DNA extraction from formalin-fixed paraffin-embedded tissues, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP), ordinary silver stain were used to study MSI and LOH of locus D17S396. Envision immunohistochemistry and Leica-Qwin computer imaging techniques were used to assess the expression of nm23H1 gene. In our experiments, the frequency of heredity instability of malignant ovarian tumors was 40%, which is higher than that of borderline ovarian tumors, while there were no heredity instability occurred in benign ovarian tumors and normal ovarian tissue. Among 25 epithelial ovarian carcinomas, the frequency of LOH in lymph node metastasis cases (66.67%) was significantly higher than those without metastasis (10.53%). Moreover, the frequency of LOH was higher in FIGO stage III and IV than in stage I and II. However, the frequency of MSI showed no correlation with any clinicopathologic characteristics. The positive frequency of nm23H1 protein in the ovarian epithelial carcinoma and borderline tumors were 56.00% and 57.14%, respectively. They were both higher than those of the benign tumors and normal ovarian tissue. In the epithelial ovarian carcinomas, the positive frequency of nm23H1 protein in lymph node metastasis case was significantly lower than those without metastasis. FIGO stage III and IV also exhibited lower positive frequency of nm23H1 protein compared with stage I and II. Furthermore, there was no difference in nm23H1 protein expression intensity analyzed by computer imaging. In the epithelial ovarian carcinomas, the positive frequency of nm23H1 protein in LOH positive group was 0.00%, which is lower than that of LOH negative group (P0.01). The results indicated that the heredity instability of nm23H1 gene might be implicated in pathogenesis and progression of epithelial ovarian tumor. The occurrence of LOH might be the molecule marker of the deteriorism of ovarian tissue. Both MSI and LOH of nm23H1 gene controlled development of the epithelial ovarian tumor independently in different paths. LOH could inhibit the expression of nm23H1 in local tissue of the epithelial ovarian carcinoma, which endowed it with high aggressive and poor prognosis. Increasing the amount of nm23H1 protein expression could effectively restrain metastasis of the ovarian epithelial carcinoma and improve prognosis of patients.

To investigate microsatellite instability (MSI) and loss of heterozygosity (LOH) of locus D17S396, D17S579 and D17S855, and their effect on the expression of nm23H1 and BRCA1 of gastric cancer, which would provide experimental basis for clinical treatment and prognosis analysis of gastric cancer. DNA was extracted from paraffin-embedded materials. Polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) was used to analyze MSI and LOH. Expression of nm23H, and BRCA1 was detected by Envision immuno-histochemistry and Leica-Qwin computer imaging techniques. In the forty cases of gastric cancer, the frequency of MSI, LOH and nm23H1 protein were 20.00%, 17.50% and 55.00% respectively at locus D17S396, while at locus D17S579, the frequency of MSI, LOH and BRCA1 protein were 22.50%, 15.00% and 37.50% respectively; at locus D17S855, the frequency of MSI, LOH and BRCA1 of thirty-seven cases were 18.92%, 18.92%, 37.84% respectively. In tumor node metastasis (TNM) staging, at locus D17S396, D17S579 and D17S855, MSI in stages I + II appeared more frequently than that in stages III + IV, while LOH appeared the contrary tendency. In the group of metastasis of gastric cancer, MSI had a less frequency (5.00%) than that with no metastasis (35.00%, P0.05) at locus D17S396, but LOH appeared more frequently (30.00%) than that with no metastasis (5.00%, P0.05). At locus D17S579, MSI had an increasing tendency with the degree of tumor differentiation (50.00% in high differentiation cases, 20.00% in middle differentiation cases, and 0% in low differentiation cases, P0.05). The frequency of nm23H1 and BRCA1 protein in stages TNM I + II was higher than that in stages TNM III + IV; and that in higher differentiation cases was higher than in poor differentiation cases. The frequency of nm23H1 protein in the group of metastasis (30.00%) was less than that with no metastasis significantly (80.00%, P0.01). The frequency of nm23H1 protein in the group positive to MSI (87.50%) was higher than that in the group negative to MSI (46.88%, P0.05). However, nm23H1 protein in group positive to LOH (14.29%) was lower than that in the group negative to LOH (63.64%, P0.05). The frequency of BRCA1 protein in the group positive to MSI (66.67%) was more than that in the group negative to MSI (29.03%, P0.05). The results of experiments indicate that MSI and LOH may separately control the development of sporadic colon cancer with different pathways. MSI may be an early period molecule marker for sporadic colon cancer, enhanced expression of nm23H1 protein can effectively inhibit colon cancer metastasis and improve prognosis of sporadic colon cancer patients. By comparison, LOH mostly arises in the late period of sporadic colon cancer and endows a high aggressive and poor prognostic phenotype. nm23H1 protein could effectively restrain gastric cancer metastasis and development; and BRCA1 protein could restain tumor from becoming lower differentiation.

The aim of this study was to evaluate the outcome of treatment of pregnancy--induced hypertension (PIH) in rats by Ligustrazine collaborated with magnesium sulfate. PIH rat models were induced with Nomega-nitro-L-arginine methyl ester (L-NAME) infusing at 7 mg/kg per day via caudal vein for four days, then treated with Ligustrazine, magnesium sulfate, or both for three days. Rat blood pressure level was measured by the tail-cuff method, 24 hours urine protein was also assayed. The blood pressure and urine proteins of grouped PIH rats were recorded before the start and after the termination of therapy. The body length and the weight of fetal rats, the weight of placentals from pregnant rats were measured. The placental tissues, livers, kidneys of rats were investigated with integrated methods such as histopathologic observation with light microscopy, ultrastructural observation with transmission electron microscopy. L-NAME administration in pregnancy rats during the late pregnancy period had resulted in an rise of blood pressure, an increasing of urine protein, death rate of rat fetus, incidence of teratogenesis, and so on. Three groups of PIH rats treated with single magnesium sulfate, Ligustrazine, or Ligustrazine combined with magnesium sulfate showed an obvious dropping of the proteinuria, decompression of blood pressure (p0.01, p0.001), especially the treatment efficacy in the group of Ligustrazine combined with magnesium sulfate was more significant effective than other two groups (p0.01, p0.001). The treatment with both Ligustrazine and magnesium sulfate could increase the body length of newly born rats, the body weight of tomites and the placental weight, furthermore, reduce the rate of the teratosis of hindlimb-shortness (p0.001). There were diffuse focal necrosis areas in the livers of PIH rats, their glomerular basement membrane had thickened extensively, the glomerular endothelium had swelled, extensive edema in the epithelia of renal tubule was demonstrated. The decidua and basal zone of the placentae of PIH rats all thickened, the microvilli of trophoblasts decreased. After treatment with ligustrazine especially with both Ligustrazine and magnesium sulfate, the necrosis of hepatocytes disappeared. The thickening of glomerular basement membrane in the group of ligustrazine or both Ligustrazine and magnesium sulfate treatment reduced; Moreover in the latter group the morphology of glomerular endothelium essentially recovered, the edema in cytoplasms of renal tubular epithelium reduced. The placental lesions were also relieved. The present results indicated the therapeutic effect by Ligustrazine collaborated with magnesium sulfate was better than a single use of Ligustrazine or magnesium sulfate. There were pathological alteration involved ischemia and anoxic in the placental tissues of PIH rats, resembled the placental pathological alteration of the human cases with PIH. The treatment with ligustrazine, and especially both Ligustrazine and magnesium sulfate in PIH rats could obviously relieve the lesions in lives, kidneys and placentae.

Classical studies have demonstrated that the visual centers in primates consist of cortical areas V1, V2 and V4 and their branches. However, nothing is known about how these visual areas change in postnatal development. In the present studies, therefore, pigs aged 2, 4, and 6 months old, were stimulated visually with a colored checker board and the active sites in the cortex, cerebellum and brainstem recorded using functional magnetic resonance imaging (fMRI). In pigs aged 2 months old, visual stimulation induced an increase in activation of sites in the V2 and V4 cortical areas, as well as in the areas of the inferior aspect of the parietal and middle aspect of the temporal cortices, but not in the medial and caudal occipital cortex (V1 area). At 4 months old, the V1 area was also activated, and by 6 months old, an inferior sector in the prefrontal cortex was also activated. As the pigs aged, functional active sites were further demonstrated in the cerebellum and the brainstem, which probably had to do with action memory, and the control of the ocular muscles, respectively. It is concluded that the visual pathway of the pig mainly involves cortical areas that mature at 6 months of age.

To study the cell signal transduction mechanism of nitric oxide (NO) on the peritoneal lymphatic stomata and lymph drainage in the rat, cGMP content were measured by a commercially available radioimmunoassay kit, and the [Ca(2+)](i) were observed by a confocal laser scanning microscope in the cultured peritoneal mesothelial cell. Animal experiment was practiced to study the effect of NO-cGMP-Ca(2+) pathway on the lymphatic stomata and lymph absorption. The results showed that: (1) Sper/NO increased cGMP of the rat peritoneal mesothelial cell (RPMC) in a dose-dependent manner (P0.01) compared to the control group. This effect was blocked by 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one (ODQ) (P0.05), a specific inhibitor of soluble guanylyl cyclase (sGC). The level of [Ca(2+)](i) in single RPMC decreased by adding Sper/NO (P0.05). Pretreatment with ODQ for 10 min blocked the Sper/NO-induced decrease in [Ca(2+)](i). L-typed calcium channel blocker nifedipine induced an immediate and marked decrease in [Ca(2+)](i) (P0.05).. After [Ca(2+)](i) reached a balance again, adding Sper/NO could not change [Ca(2+)](i) (P0.05). (2) Sper/NO increased the area of the stomata (P0.01) and the quantity of the tracer in a dose-dependent manner (P0.05) compared to the control group. Pretreatment with ODQ significantly inhibited Sper/NO-induced change of lymphatic stomata and lymph drainage (P0.01). Nifedipine increased the opening area of the lymphatic stomata (P0.01) and the concentration of absorbed trypan blue of the diaphragm (P0.05). Sper/NO could not make a further change in the samples pretreated by nifedipine (P0.05). The results indicate that NO can decrease [Ca(2+)](i) in the RPMC through the NO-cGMP pathway. This procession is related with the L- type voltage-gated Ca(2+) channel. NO enlarges the opening area of the lymphatic stomata and enhances the lymph drainage of tracer by NO-cGMP-[Ca(2+)](i) pathway.

Developmental changes in brain activation after pain stimulation and after passive movement of the hind paw were assessed by functional magnetic resonance imaging (fMRI) in pigs of postnatal ages 2, 4 and 6 months. Response patterns were correlated with histological maturation parameters. At 2 months, fMRI failed to detect brain activation after pain stimulation and revealed weak, but widespread activation after passive movement. At 4 months, strong reaction of numerous cortical areas on the contralateral side was seen after pain stimulation. Following passive movement, activation was weaker but more widespread, and the brainstem was also involved. By 6 months, cortical activation became more restricted to the contralateral sensory cortex and brainstem after pain stimulation and to the contralateral sensory and ipsilateral premotor and motor cortices after passive movement. Neocortical synaptophysin immunoreaction increased significantly between 2 and 4 months and slightly decreased by 6 months. The density of GABA-immunoreactive neurons and fibers significantly increased, reaching a maximum at 6 months. Our studies indicate that remodeling of synapses and development of inhibitory GABA neurons last until 6 months postnatally, when the fMRI response of the pig's brain also attains its mature adult pattern.

To investigate the relationship between mutations of rearranged during transfection (RET) proto-oncogene and Chinese patients with Hirschsprung's disease (HD), and to elucidate the genetic mechanism of familial HD patient at the molecular level.Genomic DNA was extracted from venous blood of probands and their relatives in two genealogies. Polymerase chain reaction (PCR) products, which were amplified using specific primers (RET, exons 11, 13, 15 and 17), were electrophoresed to analyze the single-strand conformational polymorphism (SSCP) patterns. The positive amplified products were sequenced. Forty-eight sporadic HD patients and 30 normal children were screened for mutations of RET proto-oncogene simultaneously.Three cases with HD in one family were found to have a G heterozygous insertion at nucleotide 18,974 in exon 13 of RET cDNA (18,974insG), which resulted in a frameshift mutation. In another family, a heterozygosity for T to G transition at nucleotide 18,888 in the same exon which resulted in a synonymous mutation of Leu at codon 745 was detected in the proband and his father. Eight RET mutations were confirmed in 48 sporadic HD patients.Mutations of RET proto-oncogene may play an important role in the pathogenesis of Chinese patients with HD. Detection of mutated RET proto-oncogene carriers may be used for genetic counseling of potential risk for HD in the affected families.

To investigate the apoptosis of Hep-A cells induced by hyperthermia combined with Nitric Oxide donor (Isosorbide dinitrate, ISDN) and its mechanism. The inhibitory effect on the growth of Hep-A cells was measured by MTT assay. Apoptosis of Hep-A cells was observed by electron microscopy and flow cytometry. The levels of Bcl-2 were detected with Western blot assay. It showed stronger antiproliferative ability in three experimental groups than that in control, and hyperthermia combined with ISDN group had better inhibitory effect than other groups (p0.05). With electron microscopy, marked changes of cell apoptosis were observed, including microvilli disappearance or reduction, cell shrinkage, chromatin condensation or margination and the presence of "apoptosis bodies". The apoptotic ratio induced by hyperthermia and ISDN group was higher than other groups, furthermore, the levels of Bcl-2 were decreased in three experimental groups. The present study indicated that hyperthermia combined with ISDN could induce apoptosis of Hep-A cells and be more effective than either hyperthermia or ISDN, which may be related to expression decreased Bcl-2.

In 16 human specimens the mesothelial cells of the diaphragmatic peritoneum and the pelvic parietal peritoneum were observed by transmission electron microscopy and freeze fracture replica. There were two types of cells i. e. the vesicle-containing cells and ER-cells. The former were found only in the diaphragmatic peritoneum and contained numerous cytoplasmic vesicles. These vesicles could be alone or in clusters. The clustered vesicles were often fused to form large vacuoles which opened on the free surface of the cell, into the intercellular space, or through the basement membrane, or sometimes became secretory particles on the free surface of the cells. Some vesicles were also observed in the microvilli. The latter were only found in the pelvic parietal peritoneum and contained abundant endoplasmic reticulum. The vesicle-containing cells seemed to absorb the material from the peritoneal cavity. The ER-cells contained abundant endoplasmic reticulum and Golgi apparatus, and showed high synthetic activity, possibly related to the production of peritoneal fluid.

The human dura mater and pia mater were studied by using a scanning electron microscope and a computer image processing system (C.I.P.). The human cerebral meningeal stomata are located between the mesothelial cells of the cerebral meninges. They are round or oval in shape with diameters of 0.33-2.98 microns. The cerebral meningeal stomata are stable structures, scattered or clustered together. Their density in the dura mater is greater than in the pia mater (P0.01), and they are regularly distributed. The statistical analysis showed that the stomata diameter and distribution density in the dura mater are 1.34 microns and 381.55/0.1 mm2; while in the pia mater they are 0.88 micron and 195.06/0.1 mm2 respectively. The cerebral meningeal stomata are probably part of the cerebral prelymphatic capillary system, which undertakes the cerebral lymph drainage because there are no lymphatic vessels in the brain although yet there is lymph drainage. Thus, we believe that the cerebral meningeal stomata are involved in maintaining the physiological function of the brain as part of the cerebrospinal fluid (CSF) which absorbs the cerebral interstitial fluid.

To determine the effect of the DNA repair gene XPD Arg156Arg polymorphism on the risk of lung cancer in a North-Eastern Chinese population, a hospital-based case-control study was designed consisting of 149 newly diagnosis subjects with lung cancer and 137 cancer-free control subjects matched on age (±3 years), gender and ethnicity. In the whole study group, XPD Arg156Arg was not associated with risk of lung cancer. In stratified analyses, the variant A-allele of XPD Arg156Arg was associated with increased risk of adenocarcinoma of lung (AA/AC versus CC; adjusted OR=1.65; 95% CI=1.09–2.50) ( P =0.02). Furthermore, the presence of one or two variant A-alleles was associated with increased risk for lung cancer (OR=2.49; 95% CI=1.10–5.64) ( P =0.03) and adenocarcinoma of lung (OR=5.60; 95% CI=1.52–20.56) ( P =0.005) among never-smokers only. These results suggest a possible gene–environment interaction. This is the first study to report a significant association of XPD Arg156Arg with risk of lung cancer.

The bcl-2 protein, which widely expressed in the developing central and peripheral nervous systems, has the functional role of blocking apoptosis. The purpose of this study was to map bcl-2 expression in the human enteric nervous system and investigate the value of bcl-2 immunohistochemical method in the diagnosis of Hirschsprung's disease (HD), as this has not previously been done. Rectal specimens were obtained from definitive operation of 20 patients with HD. Specimens were analyzed with immunohistochemical methods, using antibodies against bcl-2. The bcl-2 protein was expressed in myenteric and submucous neurons in normal adult and HD expand segment, but no bcl-2 immunoreactive enteric neurons was revealed in the narrow segment. And nerve fibers of the enteric plexuses that were bcl-2 immunoreactive were few in all examined specimens. From the conclusion, expression of bcl-2 is displayed in enteric neurons and immunohistochemical analysis of bcl-2 may also be valuable for identification of the enteric neurons in HD.

Minipigs, 2, 4, 6 months old, were used to evaluate the relationship between myelination in the fiber tracts of the central nervous system (CNS) of this animal during development. Histological results showed an increased density of the myelinated fibers as well as branching of these fibers in the areas studied, including the cortical white matter, olfactory tract, the corticospinal tract, the fasciculus cuneatus and the spinal V nucleus from 2 to 6 months old. By 6 months, the pig was sexually matured. Concomitantly, there was an increase in high signal-intensity regions (sites) in the magnetic resonance T(1)-weighted images as myelination progressed. There is a good correlation between the histologically observed progress of myelination and the T(1)-weighted images in the development of the CNS of the pig.

The inhibition effects of Sargassum fusiforme Polysaccharides (SFPS) on proliferation of human HL-60 promyeloid leukemia cells were measured by MTT assay. SFPS-induced apoptosis in HL-60 cells was observed by electron microscopy, flow cytometry and DNA electrophoresis. The results revealed that SFPS exhibited antiproliferative activity which depended on dosage and time. After incubation for 24, 36, 48 and 72 h, SFPS had growth inhibitory effect on HL-60 cells with IC50 of 390, 362, 402 and 421 mg/L. A ladder-like pattern of DNA fragmentation was demonstrated on agarose gel electrophoresis after HL-60 cells induced by SFPS at a dose of 300 mg/L and 500 mg/L. Morphological examination by electron microscopy showed cells with few microvilli on the surface, chromatin condensation and margination, cell shrinkage and the presence of "apoptosis bodies". The DNA content analysis of FCM showed pre-G1 cells and increased G2 period ratio, in addition, it demonstrated that apoptosis induced by SFPS was dose-and time-dependent. In conclusion, the antitumor effect of SFPS is relative to cell apoptosis and the blocking of G2/M period cells.

Techniques such as DNA extraction from paraffin-embedded tissues, polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP), ordinary silver stain, Envision immunohistochemistry and Leica-Qwin computer imaging techniques were used to study microsatellite instability (MSI) and loss of heterozygosity (LOH) of locus D17S396 at the 17th chromosome of Chinese patients and their influence on the expression of gene nm23H1, and to clarify the relationship between the genetic instability of gene nm23H1 and the development of colon cancer, which may provide experimental basis for clinical treatment. In our experiments, the frequency of MSI, LOH and nm23H1 protein reacted positive of 30 cases of colon cancer were 26.67%, 20.00% and 53.33% respectively. In tumor node metastasis (TNM) staging, the positive frequency of MSI (43.75%) and nm23H1 protein (81.25%) in stage I + II were more than those (MSI 7.14%, p0.05 and nm23H1 21.43%, p0.01) in stage III + IV, while the frequency of LOH (35.71%), which had a rising trend along with the Duke's staging increasing, was higher than that of LOH (6.25%, p0.05) in stage I + II. The positive frequency of nm23H1 protein in the group of tubular adenocarcinoma (60.00%) was distinctively higher than that in the group of mucoid adenocarcinoma (20.00%, p0.01), showing a rising trend along with the increase of the differentiation degree of tubular adenocarcinoma. Furthermore, the positive frequency of nm23H1 protein in MSI positive group was also higher than MSI negative group (p0.05). And there was no difference in nm23H1 protein expression analyzed by computer imaging techniques. The results of experiments indicated that both MSI and LOH controlled the development of sporadic colon cancer independently in different paths. LOH occurred mostly in the late period of sporadic colon cancer and endowed with it a high aggressive and poor prognosis. In contrast, MSI was an early period molecule marker of sporadic colon cancer. Increasing the amount of nm23H1 protein expression could effectively restrain colon cancer metastasis and improved prognosis of sporadic colon cancer patients.

This study aimed to investigate the effects of environmental pollutant sulfite and inorganic mercury on the peritoneal macrophages (PM phi) of mice and further explore the significance of macrophage as a biomarker in biological monitoring. PM phi were taken from the peritoneal cavity of mice and cultivated with Na2SO3 and HgCl2 respectively in vitro, and then morphologic changes were detected under light microscope and electron microscope. Also the function of PM phi was detected in yielding nitric oxide (NO), reducing MTT and phagocytizing. After affected by the pollutants, marked morphologic changes of PM phi were observed, and NO reducing, cell vigor and phagocytizing of PM phi were obviously inhibited (p0.05 or p0.01). And some PM phi were necrotic caused by HgCl2 at high concentration(10(-4) mol/L). The results suggested that PM phi of mice was damaged distinctly by using Na2SO3 and HgCl2 which showed sulfite and inorganic mercury directly impacted on non-special defensive function of macrophages. Meanwhile the experiment indicated that macrophage could be used as an effective biomarker in biological monitoring of environmental contamination.

The aim of the present study was to explore the role of vascular endothelial growth factor-C (VEGF-C) in the process of angiogenesis, lymphangiogenesis and lymphatic metastasis in epithelial ovarian tumors. In situ hybridization and immunohistochemical staining for VEGF-C were performed in 30 epithelial ovarian carcinomas, 9 borderline tumors and 26 benign cystadenomas. Endothelial cells were immunostained with anti-VEGFR-3 pAb and anti-CD31 mAb, and VEGFR-3 positive vessels and microvessel density (MVD) were assessed by image analysis. VEGF-C mRNA and protein expression in ovarian epithelial carcinomas were significantly higher than that in borderline tumors and benign cystadenomas (p0.05 or p0.01). In ovarian epithelial carcinomas, VEGF-C protein expression, VEGFR-3 positive vessels and MVD were significantly higher in the cases of clinical stage III-IV and with lymphatic metastasis than those of clinical stage I-II and without lymphatic metastasis respectively (p0.05 or p0.01), VEGFR-3 positive vessels and MVD was significantly higher in the VEGF-C protein positive tumors than negative tumors (p0.05), VEGFR-3 positive vessels was significantly correlated with MVD(p0.01). These data suggest that VEGF-C might play a role in lymphatic metastasis via lymphangiogenesis and angiogenesis in epithelial ovarian carcinomas, and VEGF-C could be used as a biologic marker of metastasis in ovarian epithelial carcinomas.

To investigate the cytotoxic effect of extracts of Trichosanthes kirilowi (TK) root on Hela cells in vitro and its probable anti-tumor mechanism.The cytotoxic effect in vitro on the growth of Hela cells was evaluated by microculture tetrazolium assay (MTT). Cell ultrastructural changes were examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), and DNA agarose electrophoresis was performed to determine apoptosis and biochemical changes of Hela cells.Exposure of Hela cells to TK extracts for 24-48 hrs resulted in a cell growth arrest, which showed in a time- and dose-dependent manner (r0.880, P0.01). With SEM and TEM, marked changes were observed, including microvilli disappearance or reduction, cell membrane vesiculation, cell shrinkage, condensation of chromosomes and apoptotic bodies with complete membrane. Besides, the apoptosis of Hela cells was confirmed by typical DNA ladder formation on gel electrophoresis.Extracts of TK has a marked anti-tumor activity and could induce apoptosis of Hela cells.

To clarify the pathogenesis of Hirschsprung's disease (HD) on molecular level and to reveal the relationship between the RET proto-oncogene and Chinese patients with HD, the mutations of the RET proto-oncogene were studied in 50 Chinese patients with HD and 30 normal children with an obstipation as control. Genomic DNA was extracted from venous blood of the HD patients and the normal children. Polymerase chain reaction (PCR) products, which were amplified using specific primers (RET; exon13), were electrophoresised to analyze the single-strand conformational polymorphism (SSCP) patterns. DNA sequences of exon 13 of the RET proto-oncogene were determined in patients showing abnormal SSCP bands. And then the familial tracking research was done for the patients with Exon 13 mutation. In 50 HD patients, seven cases were found with apparently abnormal SSCP bands. And three point mutations were proved by DNA sequencing. In the parents of seven HD patients with Exon 13 mutation, two patients' fathers had the same mutation as their children. The results suggest that the mutation of the RET proto-oncogene may have a high freqency in Chinese patients with HD, especially the heterozygous point mutation. And HD has the heredity tendency.

To investigate the expression of vascular endothelial growth factor-C (VEGF-C) mRNA and CD 31 in esophageal squamous cell carcinoma (ESCC) and its promotion of lymphatic metastasis. The expression of VEGF-C mRNA was examined in 43 ESCC by in situ hybridization. Intratumoral microvessel density (MVD) was assessed by immunostaining endothelial cells, using anti-CD31 antibody. The positive rate of VEGF-C mRNA expression was 41.86%. The average rank of MVD was 76.36 +/- 20.30/mm2. VEGF-C mRNA expression correlated significantly with lymph node metastasis, TNM stage and depth of invasion (p0.05 or p0.01) in statistic, but not with histological grade (differentiation) (p0.05). MVD correlated significantly with lymph node metastasis and TNM stage (p0.05) in statistic, but not with depth of invasion and histological grade (differentiation) (p0.05). MVD was significant higher in the VEGF-C positive tumors than negative tumors (p0.05). The present study indicated that VEGF-C might play a role in lympatic metastasis via lymphangiogenesis and angiogenesis in ESCC.

The aim of the experiment is to establish two murine hepatocarcinoma (HepA cell line) substrains with different lymphatic metastatic ability and compare their biological characteristics. Two murine hepatocarcinoma substrains, HepA-H and HepA-L, were established by the cell cloning and the selection way of lymph node metastasis. After tumor cells were injected subcutaneously to foot pad of NIH mice, the popliteal lymph node metastatic rates of HepA-H and HepA-L were 83.3% and 16.7% respectively, which has statistical significance (p0.01). There were no metastatic lesions were found in other organs, such as lung, liver, spleen and kidney. In vitro, HepA-H showed a higher ability in growth, migration and adhesion than that of HepA-L. The establishment of two murine hepatocarcinoma substrains provides a useful animal model for the study of mechanism of the lymphatic metastasis.

To set up a nontraumatic and convenient mouse model of severe acute pancreatitis (SAP). Caerulein(Cn) was injected the mice intraperitonealy with lipopolysaccharide(LPS). Serum amylase and pancreas weight were measured in experiment. The pathological changes of pancreas and other organs were observed under light microscope. The ultrastructure of acini were observed under transmission electron microscope (TEM). Serum NO concentration were measured and the SOD and MDA in pancreas were examined. The results in Cn + LPS group were showed that serum amylase, NO concentration and pancreas weight were increased, SOD deduced and MDA increased. Severe edema, inflammation infiltration, necrosis and different extent of hemorrhage were showed. The acini were damaged severely. And the lesion of other organs were also happened. In Cn group, there were only pancreatic interstitial edema but no parenchmal necrosis or hemorrhage, and the other organs were normal. In LPS group, pancreas were almost normal and the organs besides pancreas were only showed light inflammation infiltration. The SAP mouse model induced by caerulein plus LPS has the same pathological characteristics of human SAP, which can be used in human SAP research. The unbalance of oxygen free radical release-elimination and oxidation-antioxidation mechanisms might be involved in the pathogenesis of mouse model of severe acute pancreatitis induced by intraperitoneal injection of caerulein plus LPS.

In order to study the influence of pregnancy and hormone on the lymphatic stomata of guinea pig's ovary bursa, the lymphatic stomata of ovary bursa were observed during pregnancy and by using exogenous hormone, including serum gonadotrophin (PMSG) combined with human chorionic gonadotropin (HCG) to promote ovulation and androgen to inhibit the ovary. Then the lymphatic stomata in the inner layer of ovary bursa and their absorption functions were observed with scanning electron microscope (SEM) and trypan blue as a tracer respectively. The lymphatic stomata were also counted by the Elescope computer image processing system. We found that the quantity of tracer absorbed by the lymphatic stomata of ovary bursa in pregnant group was more than that in ovulation-promoted group, and least in the androgen-treated group, which had statistical significance between each couples of them (P0.05). The opening area of the lymphatic stomata in pregnant, ovulation-promoted and androgen-treated groups were 189.9 +/- 48.7 micron 2/1000 micron 2, 104.4 +/- 31.2 micron 2/1000 micron 2 and 40.5 +/- 18.7 micron 2/1000 micron 2 respectively, which had remarkable statistical difference between each couples (P0.01). Under SEM observation, the secretory granules in ciliated columnar epithelium were less in pregnant group than the other two, while both the secretory granules and the cilium were most active in ovulation-promoted group. The results indicated that both the openings and absorption functions of the lymphatic stomata in guinea pig's ovary bursa could be affected by pregnancy and exogenous homone. There existed relationship between ovary function and the regulation of the lymphatic stomata in ovary bursa.

OBJECTIVE: To investigate the development of the enteric nervous system(ENS) in the middle stage(the 4th approximate, equals 6th months) of human embryogenesis. METHODS: A histologic evaluation of the clonic enteric nervous system was done using NADPHd histochemistry, PAP immunohistochemistry, with anti-PGP 9.5 and anti-S-100 protein. RESULTS: During this stage of embryology three things were noted. (1)The nerves in the myenteric layer increased markedly. This was shown by the PGP 9.5 immunoreactive nerves spreading out and the S-100 protein immunoreactive nerves forming a "bamboo basket"pattern. (2)The whole myenteric colon showed nitrigeric nerves paralleling the growth of the myofibers in the circular muscle layer. Nitrigeric perikara were rarely found in the extrinsic submucosal layer.(3) In the whole-mounted preparations reactive nerves formed the complex nerve net in the myenteric layer. CONCLUSION: The middle stage of embryogensis is very important to the development of the colon ENS.

To study the effect of Angelica sinensis, Salvia miltiorrhiza and Ligustrazine on function of peritoneal macrophages during peritoneal dialysis.Peritoneal macrophages of mice were cultured in culture medium (control), peritoneal dialysate (PD), drugs contained PD containing Angelica, Salvia and Ligustrazine combined (PD-ASL) or separated (PD-A, PD-S, PD-L) with concentration of 2 micrograms/ml, 10 micrograms/ml and 100 micrograms/ml, separately for 24 hrs. The nitric oxide (NO) content, methyl thiazolyl tetrazolium (MTT) reducing capacity (MTT-RC) and phagocytosis capacity of macrophages were determined and compared.NO content and MTT-RC of macrophages cultured in PD group were significantly lower than those of the control (P0.01), as compared with those in drug contained PD groups, the NO content in the PD-L group and the MTT-RC in the PD-ASL group were higher significantly (P0.01). The phagocytosis capacity and NO content in the PD-ASL group were raised along with the increased concentration of drug in PD.Administering Chinese herbal medicine during peritoneal dialysis has important significance in improving the defense function of peritoneal macrophages, reducing the incidence of peritonitis and enhancing the therapeutic effect of peritoneal dialysis.

OBJECTIVE: To investigate the influence of commercial peritoneal dialysate (CDS) on function of macrophage. METHODS: Cultured peritoneal macrophages were divided into two experimental groups and their controls.(1) Macrophages were cultured in conditioned culture medium containing 50%CDS (0.139 mol/L glucose) for 24 h. (2)Macrophage were exposed to CDS containing 0.139mol/L glucose for 10, 30 and 60 min respectively, and then cultured in CDS-free medium for 24 h. The nitric oxide (NO) production and MTT in two groups were measured. In each control group, CDS was replaced by same amount of culture medium. RESULTS: NO production and MTT reduction ability (related to cell viability) of experimental groups were remarkably lower than those of controls and the NO production and MTT reduction in 60min CDS-exposed group were lower than that of 10 min and 30 min CDS-exposed group. CONCLUSION: Dialysate may have detrimental effects on viability and other function of macrophage and these effects may related to time length of CDS exposure.

Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 – August 2, 2012.

Aim To observe the regulatory effect of Chinese herbal medicine on peritoneal lymphatic stomata and its significance in treating ascites in liver fibrosis model mice. Methods Two Chinese herbal composite prescriptions were used separately to treat the carbon tetrachloride-induced mouse model of liver fibrosis. The histo-pathologic changes of the liver sections (HE and VG stainings) were observed. The peritoneal lymphatic stomata was detected by scanning electron microscopy and computer image processing. The changes of urinary volume and sodium ion concentration were measured. Results In the model group, lots of fibrous tissue formed in liver and extended into the hepatic lobules to separate them incompletely. In the treated and prevention groups, the histo-pathologic changes of liver was rather milder, only showed much less fibrous tissue proliferation in the hepatic lobules. The peritoneal lymphatic stomata enlarged with increased density in the experimental groups (diameter: PA, 3.07 +/- 0.69 microm; PB, 2.82 +/- 0.37 microm; TA, 3.25 +/- 0.82 microm and TB, 2.82 +/- 0.56 microm; density: PA, 7.11 +/- 1.90 stomata.1000 microm(-2); PB, 8.76 +/- 1.45 stomata.1000 microm(-2); TA, 6.55 +/- 1.44 stomata.1000 microm(-2)and TB, 8.76+/-1.79 stomata.1000 microm(-2)), as compared with the model group (diameter: 2.00+/-0.52 microm density: 4.45+/-1.05 stomata.1000 microm(-2)). After treatment, the urinary volume and sodium ion excretion increased in the experimental groups (PA, 231.28+/-41.09 mmol.L(-1); PB, 171.69 +/- 27.48 mmol.L(-1) and TA, 231.44 +/- 34.12 mmol.L(-1)), which were significantly different with those in the model group (129.33 +/- 36.75 mmol.L(-1)). Conclusion Chinese herbal medicine has marked effects in alleviating liver fibrosis, regulating peritoneal lymphatic stomata, improving the drainage of ascites from peritoneal cavity and causing increase of urinary volume and sodium ion excretion to reduce the water and sodium retention, and thus have favorable therapeutic effect in treating ascites.

AIM: To study the mechanism of Chinese herbal medicine (CHM, the prescription consists of Radix Salviae Miltiorrhizae, Radix Codonopsitis Pilosulae, Rhizoma Atractylodis Alba and Rhizoma Alismatis, Leonurus Heterophyllus Sweet, etc) on the regulation of the peritoneal lymphatic stomata and the ascites drainage. METHODS: The mouse model of live fibrosis was established with the application of intragastric installations of carbon tetrachloride once every three days; scanning electron microscope and computer image processing were used to detect the area and the distributive density of the peritoneal lymphatic stomata; and the concentrations of urinary ion and NO in the serum were analyzed in the experiment. RESULTS: Two different doses of CHM could significantly increase the area of the peritoneal lymphatic stomata, promote its distributive density and enhance the drainage of urinary ion such as sodium, potassium and chlorine. Meanwhile, the NO concentration of two different doses of CHM groups was 133.52 ± 23.57 μmol/L, and 137.2 ± 26.79 μmol/L respectively. In comparison with the control group and model groups (48.36 ± 6.83 μmol/L, and 35.22 ± 8.94 μmol/L, P < 0.01), there existed significantly marked difference, this made it clear that Chinese herbal medicine could induce high endogenous NO concentration. The effect of Chinese herbal medicine on the peritoneal lymphatic stomata and the drainage of urinary ion was altered by adding NO donor(sodium nitropurruside, SNP) or NO synthase (NOS) inhibitor (N(G)-monomethyl-L-arginine, L-NMMA) to the peritoneal cavity. CONCLUSION: There existed correlations between high NO concentration and enlargement of the peritoneal lymphatic stomata, which result in enhanced drainage of ascites. These data supported the hypothesis that Chinese herbal medicine could regulate the peritoneal lymphatic stomata by accelerating the synthesis and release of endogenous NO.

AIM:To study the development of gastroenteric nervous system in trisomy 16 mouse embryos.The gastroenteric nervous system in trisomy 16 mice and their normal littermates, serving as controls from embryonic days 13 to 18 (ED13-18) was identified by using primary antibody against protein gene product (PGP) 9.5.METHODS:Trisomy 16 mouse breeding and trisomy 16 mouse embryos were identified from their normal littermates by chromosome examination; PGP 9.5 immunohistochemical stainning.RESULTS:In normal littermates embryos, the precursor cells from the neural crest migrated into stomach and intestine at ED 13 and ED 14 respectively.Numerous nervous processes connected to each other and formed early nervous networks at ED 14 stomach and ED 15 intestine. Original ganglia in the muscular nervous plexus of the stomach appeared at ED15 with very simple arrangement. At ED 16 the early developed myenteric nervous plexuses were regularly found in the stomach and intestine respectively. In both stomach and intestine, the development of submucosal nervous plexuses were finished at ED17. However, the myenteric nervous plexus and the internal and external submucosal nervous plexuses were differentiated only in the stomach at ED 18.In comparison with the normal littermates, stomach and intestine nervous system developed much slower in trisomy 16 mice. Their immature neurons did not appear in the stomach and intestine until ED 14 and ED 15. Between ED 14 and ED 16, the gastroenteric nervous system was composed of only some scattered neurons with different distribution density and size. The development and differentiation of the gastroenteric nervous system were delayed and the myenteric nervous plexus did not appear until ED 18. There was no submucosal nervous plexus in all stomach and intestine specimens. A semiquantitative analysis and rank sum test of the data showed that the trisomy 16 mouse embryos were markedly retarded in the gastroenteric nervous development compared with their normal littermates.CONCLUSION:Trisomy 16 mice, as an animal model for Down syndrome, has abnormality not only in several systems and organs but also in gastroenteric innervation. This report describes for the first time that the development of the gastroenteric nervous system was not only delayed but also pathological.

To establish whether cystic fibrosis transmembrane conductance regulator (CFTR) is functionally expressed in the testis, we subjected spermatogenic cells from rat testes to analysis of CFTR mRNA, protein and channel activity. CFTR mRNA was detected in the testes of mature but not immature rats using reverse transcription-polymerase chain reaction analysis. Western blot analysis performed with a CFTR specific antibody revealed immunoreactivity in the membrane extract of spermatogenic cells. Immunohistochemical studies localized CFTR in round and elongated spermatids, but not in the fully developed spermatozoa. Using a whole-cell patch clamp technique, we recorded an inward current activated by intracellular cAMP (100 micromol/l) in round spermatids. The current displayed a linear I / V relationship and was inhibited by diphenylamine-2-carboxylate (DPC), a chloride channel blocker. Transfection of the rat germ cell CFTR cDNA into human embryonic kidney (HEK) 293 cells caused the expression of a cAMP-activated chloride current with CFTR characteristics. The current was completely blocked by the antispermatogenic agents 1-(2,4-dichlorobenzyl)-indazole-3-carboxylic acid, lonidamine (500 micromol/l) and 1-(2,4-dichlorobenzyl)-indazole-3-acrylic acid, AF2785 (250 micromol/l). These results taken together provide evidence that CFTR is differentially expressed in spermatids during spermiogenesis. We speculate that CFTR may interact with aquaporin to bring about cytoplasmic volume contraction which is an essential feature of spermiogenesis.

The pool boiling heat transfer of FC-72 on a plain plate with a heating area of 15*15 mm2 in different pressure and subcooling has been studied experimentally both in normal gravity on the ground and in microgravity aboard the Chinese recoverable satellite SJ-8. A quasi-steady heating method is adopted, in which the heating voltage is controlled as an exponential function with time. Three modes of heat transfer, namely single-phase natural convection, nucleate boiling, and transition boiling, are observed, while the nucleate pool boiling are the major subject for discussion in the present paper. In normal gravity, the present data is compared with those obtained by other researchers, and a satisfactory agreement is evident and warrants reasonable confidence in the data. Compared with terrestrial experiments, the boiling curves in microgravity are considerably gentle, and CHF is no more than 40% of that in terrestrial condition. In microgravity, influences on boiling curves of the pressure and subcooling are similar with those in normal gravity, namely that the heat transfer coefficient and CHF will increase with subcooling for the same pressure, and that they will also increase with pressure for the same subcooling. Keywords: microgravity, pool boiling, subcooling, plate heater

Background It is very difficult to prevent pulmonary tuberculosis (TB) due to the lack of specific and diagnostic markers, which could lead to a high incidence of pulmonary TB. We screened the differentially expressed serum microRNAs (miRNAs) as potential biomarkers for the diagnosis of pulmonary TB. Methods In this study, serum miRNAs were screened using the Solexa sequencing method as the potential biomarkers for the diagnosis of pulmonary TB. The stem-loop quantitative reverse-transcription polymerase chain reaction (qRT-PCR) assay was used to verify differentially expressed serum miRNAs. The receiver operating characteristic (ROC) curve and logistic regression model were used to analyze the sensitivity and specificity of the single miRNA and a combination of miRNAs for diagnosis, respectively. Using the predicted target genes, we constructed the regulatory networks of miRNAs and genes that were related to pulmonary TB. Results The Solexa sequencing data showed that 91 serum miRNAs were differentially expressed in pulmonary TB patients, compared to healthy controls. Following qRT-PCR confirmation, six serum miRNAs (hsa-miR-378, hsa-miR-483-5p, hsa-miR-22, hsa-miR-29c, hsa-miR-101 and hsa-miR-320b) showed significant difference among pulmonary TB patients, healthy controls (P

Background Noninvasive and convenient biomarkers for early diagnosis of tuberculosis (TB) remain an urgent need. The aim of this study was to discover and identify potential biomarkers specific for TB. Methods The surface-enhanced laser desorption ionization time of flight mass spectrometry (SELDI-TOF MS) combined with weak cation exchange (WCX) magnetic beads was used to screen serum samples from 180 cases of TB and 211 control subjects. A classification model was established by Biomarker Pattern Software (BPS). Candidate protein biomarkers were purified by reverse phase-high performance liquid chromatography (RP-HPLC), identified by MALDI-TOF MS, LC-MS/MS and validated using enzyme-linked immunosorbent assay (ELISA). Results A total of 35 discriminating m/z peaks were detected that were related to TB (P

BACKGROUND: A large number of studies have investigated whether polymorphisms in the Toll-like receptor (TLR) genes are implicated in susceptibility to tuberculosis (TB) in different populations. However, the results are inconsistent and inconclusive. METHODS: A literature search was conducted using the PubMed, EMBASE, Medline (Ovid), ISI Web of Knowledge and Chinese National Knowledge Infrastructure (CNKI). A meta-analysis on the associations between the TLR1 G1805T, TLR2 T597C, T1350C, G2258A, and TLR6 C745T polymorphisms and TB risk was carried out by comparison using different genetic models. RESULTS: In total, 16 studies from 14 articles were included in this review. In meta-analysis, significant associations were observed between the TLR2 2258AA (AA vs. AG+AG, OR 5.82, 95% CI 1.30-26.16, P = 0.02) and TLR6 745TT (TT vs. CT+CC, OR 0.61, 95% CI 0.39-0.97, P = 0.04) polymorphisms and TB risk. In the subgroup analysis by ethnicity, Africans and American Hispanic subjects with the TLR1 1805T allele had an increased susceptibility, whereas Asian and European subjects with the TLR2 2258A allele had an increased susceptibility to TB. CONCLUSIONS: The meta-analysis indicated that TLR2 G2258A is associated with increased TB risk, especially in Asians and Europeans. TLR1 G1805T is associated with increased TB in Africans and American Hispanics. TLR6 C745T is associated with decreased TB risk. Our systematic review and meta-analysis reported an interesting preliminary conclusion, but this must be validated by future large-scale and functional studies in different populations.

The experiment on mice was carried out by injecting intraperitoneally Chinese materia medica for treating hepatocirrhosis with ascites. Observations and a quantitative analysis were carried out on the pharmacological regulation of the peritoneal stomata by using a scanning electron microscope (SEM) and a computer image processing system attached to the SEM. There was a significant increase in both the diameter (P0.05) and distribution density (P0.01) of the peritoneal stomata in the red sage root and alismatis rhizome groups, whereas the effect of poria and poria peel was not significant compared with the control group (P0.05). Our findings confirm the effect of red sage root and alismatis rhizome on the regulation of the peritoneal stomata, which can enhance the absorption of ascitic fluid, taking into consideration the absorbent function of these stomata. They indicate that the patency of peritoneal stomata can vary in response to the effect of some Chinese materia. They also suggest that the ascites is drained mainly by means of enhancing the patency of the stomata and lymphatic absorption of the stomata during the process of treatment by traditional Chinese medicine.

The epithelium of normal and ligated Fallopian tubes in nine women of reproductive age was studied by using scanning electron microscopy (SEM). The results were as follows: at a distance of 0.5 cm from the ligated scar, numerous cilia of the epithelium were twisted and adhered, the shape of the epithelial cells was irregular, and even many microvilli and cilia lost from the local epithelium. However, the microvilli and cilia of the epithelium were normal on the mucosa at 1.0 cm from the ligated scar. This may account for the continued infertility following re-anatomosis of the ligated Fallopian tubes, due to the ultrastructural changes of oviducts and inability to convey the ova, Thus, at the Fallopian tube re-anastomosis, it is recommended that the removal of the mucosa should be at least 0.5 cm away from the ligated scar so as to increase the success rate of the fertilization. On the epithelium of the hydrosalpinx, Fallopian tube stomata located among the secretory cells were found for the first time. These stomata are small openings of lymphatic capillaries with a diameter of 3.25 μm to 4.25 μm. They are likely to be connected between the lumen of the Fallopian tube and subserous lymphatic vessels, and also involved in the drainage of the hydrosalpinx. Most or all cilia were lost from the ciliated cell surface on the epithelium of the hydrosalpinx, whereas the microvilli were flourishing. Therefore, many ciliated cells turned into secretory cells. In addition, a new pathogenesis of the hydrosapinx is discussed.

Effects of melatonin on hypothalamic neurotransmitters in male mice were studied. Exogenous melatonin administered intraperitoneally significantly increased (p0.05) hypothalamic concentrations of aspartic acid and gamma-aminobutyric acid by over 29 and 50% respectively. Conversely, hypothalamic beta-endorphin concentration was significantly decreased (p0.05) 30 min after melatonin administration with doses between 5- and 100 micrograms/kg. Similarly, melatonin, at a concentration of 100 micrograms/kg, decreased (p0.05) the serotonin level in mouse hypothalamus by 46%. Melatonin, however, did not affect the concentration of hypothalamic glutamic acid over a dose range of 0.5-300 micrograms melatonin/kg. Our findings suggested that actions of pineal melatonin in animals such as inhibition on serum corticosterone levels might be mediated by the potentiation of activities of hypothalamic neurons containing gamma-aminobutyric acid and aspartic acid or by the inhibition of the beta-endorphin and serotonin hypothalamic neurons. The neurons containing glutamic acid in the hypothalamus were, however, not influenced by melatonin. Our results are in line with the suggestion that melatonin actions on adrenal corticosterone release or other endocrine secretions may be mediated by way of its actions on hypothalamic neurotransmitter activities.

AIM: To investigate the role of transforming growth factor (TGF)-β-inducible early gene 1 (TIEG1) in TGF-β-induced growth inhibition in hepatocellular carcinoma (HCC) cells. METHODS: Human hepatocyte and HCC cell lines with varied susceptibilities to TGF-β1 were tested by methylthiazoletetrazolium (MTT) assay. The expression changes of Smad2, Smad3, Smad4, Smad7, TIEG1 and TIEG2 gene following treatment with TGF-β1 in a TGF-β-sensitive hepatocyte cell line (MIHA), a TGF-β-sensitive hepatoma cell line (Hep3B) and two TGF-β-insensitive hepatoma cell lines (HepG2 and Bel7404) were examined. SiRNA targeting TIEG1 was transfected into Hep3B cells and the sensitivity of cells to TGF-β1 was examined. Overexpression of TIEG1 was induced by lentiviral-mediated transduction in TGF-β1-resistant hepatoma cell lines (Bel7404 and HepG2). MTT assay and 4’,6-Diamidino-2-phenylindole staining were used to identify cell viability and apoptosis, respectively. The expression level of stathmin was measured by reverse transcriptase polymerase chain reaction and Western-blotting analysis, and stathmin promoter activity by TIEG1 was monitored by a luciferase reporter gene system. RESULTS: TIEG1 was significantly upregulated by TGF-β1 in the TGF-β1-sensitive HCC cell line, Hep3B, but not in the resistant cell lines. The suppression of TIEG1 by siRNAs decreased the sensitivity of Hep3B cells to TGF-β1, whereas the overexpression of TIEG1 mediated growth inhibition and apoptosis in TGF-β1-resistant HCC cell lines, which resembled those of TGF-β1-sensitive HCC cells treated with TGF-β1. Our data further suggested that stathmin was a direct target of TIEG1, as stathmin was significantly downregulated by TIEG1 overexpression, and stathmin promoter activity was inhibited by TIEG1 in a dose-dependent manner. CONCLUSION: Our data suggest that transactivation of TIEG1 conferred growth inhibition of TGF-β-susceptible human HCC cells.