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Objective To analyze the effects of nicotine on the bone calcium and phosphorus content and alkaline phosphatase (ALP) activity in rat alveolar bone and mandible. Methods Twenty health male Wistar rats of five weeks of age were randomly assigned to two groups and received daily intraperitoneal injections for three months as follows: Saline solution for control group, nicotine 0.73 mg·kg-1·d-1 for experimental group. The bone calcium phosphorus content were detected by concentrated acid digestion method and the ALP activity was examined by improved Reddi method. Results Compared to the control group, bone calcium and phosphorus content was lower in the experimental group (P>0.05), ALP activity had no statistical significance(P>0.05). Bone calcium phosphorus and ALP activity in different parts had no statistical significance (P>0.05). Conclusion The nicotine reduces calcium phosphorus deposition of jaw bone, but has no obvious influence to ALP activity. [ABSTRACT FROM AUTHOR]

Microcystin-leucine arginine (MC-LR), a potentially carcinogenic toxin, is produced by Cyanobacteria such as Microcystis and Ananabacteria during water bloom. Increasing evidence demonstrated that MC-LR induces male reproductive toxicity, mainly by inducing germ cell apoptosis, destroying cell cytoskeleton, interfering with DNA damage repair pathway, and damaging blood-testicular barrier (BTB), which eventually lead to male sterility. Testicular Sertoli cells are the somatic cells that directly contact with spermatogenic cells in seminiferous tubules. They not only regulate immune response to maintain testicular immune homeostasis by secreting a variety of cytokines and immunosuppressive factors, but also provide the protective effects of spermatogenic cells by forming BTB. MC-LR induces inflammation and apoptosis of Sertoli cells, and destroys the integrity of the BTB, and then causes spermatogenesis dysfunction.

Objective: To investigate the role of inositol-requiring enzyme 1(IRE1) in autophagy of human gastric cancer cells induced by vitamin E succinate(VES)., Methods: Human gastric cancer SGC-7901 cells were cultured in vitro and divided into solvent control group(0.1% ethanol absolute), different doses(5, 10, 15 and 20 μg/mL) VES group, 4μ8C group, and VES + 4μ8C group. The endoplasmic reticulum stress-related molecules glucose regulated protein 78(GRP78) and C/EBP homologous protein(CHOP), autophagy marker microtubule associated Protein1 light chain 3(LC3), Beclin-1, unfolded protein response branching pathway Inositol-requiring enzyme 1(IRE1), X box-binding protein 1(XBP1), c-Jun n-terminal kinase(JNK) and p-JNK were detected by Western blot in the solvent control group and different doses of VES group. IRE1 was inhibited by 4μ8C. The expressions of IRE1, XBP1, JNK, p-JNK, GRP78 and CHOP were detected by Western blot, and the expressions of LC3 and Beclin-1 were detected., Results: The expression of GRP78(1.16±0.06) and CHOP(1.36±0.11) in 20 μg/mL VES group were significantly higher than those in solvent control group GRP78(0.36±0.10) and CHOP(0.48±0.05)(P<0.001). The expression of Beclin-1(1.09±0.20) and LC3-Ⅱ/LC3-Ⅰ(1.29±0.03) in 20 μg/mL VES group were significantly higher than those in solvent control group(0.27±0.07) and LC3-Ⅱ/LC3-Ⅰ(0.43±0.06)(P<0.001). The expression levels of IRE1(1.07±0.20), XBP1(1.33±0.07) and p-JNK/JNK(1.19±0.31) in 20 μg/mL VES group were significantly higher than those in the solvent control group(P<0.01). After IRE1 is inhibited: The expression level of IRE1(0.63±0.27), XBP1(0.74±0.09), p-JNK/JNK(0.35±0.04), GRP78(0.66±0.02), CHOP(0.51±0.02), LC3-Ⅱ/LC3-Ⅰ(0.72±0.01), Beclin-1(0.70±0.15) was significantly lower than that of VES group(P<0.05)., Conclusion: VES may participate in the regulation of autophagy in gastric cancer cells by upregulating IRE1 pathway.

Objective To explore the effects of D-galactose (D-gal) on barrier function of murine TM4 sertoli cells and its mechanism. Methods TM4 cells were divided into control group and 25, 50, 100, 150, 200, 250 mmol/L D-gal stimulation group. The viability of TM4 cells was determined by MTT assay. The protein expression levels of tight junction-related proteins including zonula occluden-1 (ZO-1) and occludin, adheren junction-related proteins including neural cadherin (N-cadherin), epithelial cadherin (E-cadherin) and β-catenin, gap junction-related protein connexin43 (CX43) and cytoskeleton-related protein vimentin, and MAPK signaling pathway-related proteins ERK1/2, phosphorylated ERK1/2 (p-ERK1/2), JNK, phosphorylated JNK (p-JNK), p38MAPK and phosphorylated p38MAPK (p-p38MAPK) were detected by Western blot analysis. Results Compared with the control group, the viability of TM4 cells significantly decreased when the concentration of D-gal was more than 50 mmol/L. In addition, the protein expression levels of ZO-1, occludin, N-cadherin, E-cadherin and β-catenin were significantly down-regulated in D-gal-treated group, while the protein expression levels of p-p38MAPK were significantly up-regulated. However, there were no differences in the protein expression levels of CX43, vimentin, p-ERK, ERK1/2, p-JNK and JNK between the control group and D-gal-treated groups. Conclusion D-gal can disrupt tight junction and adheren junction of TM4 cells via the activation of p38MAPK signaling pathway.

Objective: To study the effect of rapamycin on scar formation in rabbit eyes following filtering operation and explore the possible mechanism., Methods: Ninety-six healthy adult rabbits were subjected to trabeculectomy of the left eye and subsequently randomly divided into 4 groups ( n =24) for treatment with castor oil (control) or rapamycin (1%, 3%, or 5%) eye drops of the operated eyes 4 times a day. The morphology and function of the filtering blebs of the rabbits were compared at 7, 14, 21 and 28 days after the operation; at each of the time points, 6 rabbits from each group were euthanized for detection of expressions of proliferating cell nuclear antigen (PCNA) and α-smooth muscle actin (α-SMA) in the tissues in the surgical area using immunohistochemistry. Cultured rabbit subconjunctival fibroblasts (RTFSs) were treated with different concentrations of rapamycin (0.06, 0.25, 1, and 4 mg/L) and the cell apoptosis was detected using flow cytometry., Results: In the first, second and third weeks after the operation, the rate of functional follicle formation was significantly higher in the 3 rapamycin groups than in the control group ( P < 0.05), and the number of α- SMA-positive fibroblasts decreased over time in the 3 rapamycin groups. In cultured RTFSs, treatment with rapamycin at different concentrations resulted in increased apoptosis of the cells, and rapamycin above 0.25 mg/L significantly increased the cell apoptosis in a dose-dependent manner., Conclusions: Rapamycin can inhibit hyperplasia of the filtering passage tissue, helps to preserve the functional filtering blebs and prolong their life span, and induces apoptosis of RTFS.

Objective: To study the anti- scarring effect of rapamycin in rabbits receiving glaucoma filtering surgery., Methods: Ninety-six Chinchilla rabbits were randomized equally into 3 rapamycin treatment groups and one control group. All the rabbits underwent trabeculectomy, after which the rabbits in the 3 rapamycin groups were treated with eye drops containing 1%, 3%, or 5% rapamycin in the operated eyes, and those in the control groups were given castor oil 4 times a day. The intraocular pressure (IOP) and inflammatory reaction in the treated eyes were observed, and the PCNA-positive cells in the filtering bleb were detected using immunohistochemistry. RTFs isolated from the Tenon's capsule of the rabbits were cultured in vitro , and the expressions of caspase-3, caspase-8, and caspase-9 in the fibroblasts were detected after treatment with different concentrations of rapamycin., Results: The IOP was significantly lower in rapamycin-treated group than in the control group after the surgery ( P &lt; 0.05). The counts of the PCNA-positive cells were significantly lower in rapamycin-treated rabbits than in the control group ( P &lt; 0.05). Rapamycin treatment dose-dependently increased the expressions of caspase-3 and caspase- 9 at both the mRNA ( P &lt; 0.001) and protein ( P &lt; 0.001) levels without causing significant changes in the expressions of caspase-8., Conclusions: Rapamycin can inhibit excessive proliferation of the fibroblasts in the filtering bleb to reduce scar formation after glaucoma filtration surgery in rabbits. Rapamycin also increases the expressions of caspase-3 and caspase-9 to induce apoptosis of the RTFs.

Objective To establish a model of decline in secretion of senescent TM4 cells in vitro induced by D-galactose (D-gal). Methods Different concentrations of D-Gal (25, 50, 100, 150, 200 and 250 mmol/L) were used to induce TM4 cell senescence. The viability of TM4 cells was detected by MTT assay. The cell cycle was analyzed by flow cytometry. The cell morphology was observed by light microscopy and the percentage of senescent cells was observed by senescence-associated β-galactosidase (SA-β-Gal) staining. The mRNA expression levels of P21, P16, glial-derived neurotrophic factor (GDNF) and stem cell factor (SCF) were detected by reverse transcription PCR. The protein expression levels of GDNF, SCF, nuclear factor erythroid 2 like 2 (NRF2), NAD(P)H dehydrogenase [quinone] 1 (NQO-1) and heme oxygenase-1 (HO-1) were detected by Western blot analysis. Results Compared with normal control group, D-Gal stimulation significantly decreased the cell viability in a concentration-dependent manner. The arrest of D-Gal-treated cells in G1 phase of cell cycle significantly increased, while it significantly decreased in S phase, and D-Gal induced cell cycle arrest at G0/G1 phase in TM4 cells. The percentages of SA-β-Gal positive cells increased significantly. The expression levels of P21 and P16 mRNAs were significantly up-regulated. The mRNA and protein levels of GDNF and SCF-1 were significantly down-regulated. Furthermore, the expression levels of oxidative stress-related protein NRF2, HO-1 and NQO-1 were significantly reduced. Conclusion The model of declined secretion function of senescent TM4 cells induced by D-Gal we established is stable and reliable. Its mechanism may be related to the down-regulation of NRF2 signaling pathway.

Objective: To analyze the effects of nicotine on the bone calcium and phosphorus content and alkaline phosphatase (ALP) activity in rat alveolar bone and mandible., Methods: Twenty health male Wistar rats of five weeks of age were randomly assigned to two groups and received daily intraperitoneal injections for three months as follows: Saline solution for control group, nicotine 0.73 mg.kg-l.d-1 for experimental group. The bone calcium phosphorus content were detected by concentrated acid digestion method and the ALP activity was examined by improved Reddi method., Results: Compared to the control group, bone calcium and phosphorus content was lower in the experimental group (P<0.05), ALP activity had no statistical significance(P>0.05). Bone calcium phosphorus and ALP activity in different parts had no statistical significance (P>0.05)., Conclusion: The nicotine reduces calcium phosphorus deposition of jaw bone, but has no obvious influence to ALP activity.

Objective: To study the chemical constituents of the dried rhizome of Ardisia gigantifolia., Method: The 60% ethanol extract was extracted with EtOAc, and then separated and purified by column chromatography using silica gel and preparative HPLC. Their structures were identified on the basis of spectral analysis and physico-chemical properties., Result: Nine compounds were isolated and identified as 11-O-galloylbergenin (1), 11-O-syringylbergenin (2), 11-O-protocatechuoylbergenin (3), 4-O-galloylbergenin (4), 11 -O-vanilloylbergenin (5), (-) -epicatechin-3-gallate (6), stigmasterol-3-O-beta-D-glucopyranoside (7), (-) -4'-hydroxy-3-methoxyphenyl-beta-D-[6-O-(4"-hydroxy-3", 5"-dimethoxybenzoyl)] -glucopyranoside (8), and beta-sitosterol (9)., Conclusion: Compounds 3, 4 and 7 were isolsted from the genus Ardisia for the first time, while compounds 1, 2, 5 and 6 were isolated from this plant for the first time.

Objective: To clone and characterize the DNA and cDNA sequences of phenylalanine ammonia-lyase gene (PAL) from Fagopyrum dibotrys, and investigate the biological activity of the obtained PAL., Method: Using homology cloning and RT-PCR techniques, the DNA and full-length cDNA sequences of PAL gene were amplified from F. dibotrys. The obtained sequences were analyzed by bioinformatics software. The ORF of PAL gene was cloned into expression vector pET-30b(+) and transformed into Escherichia coli BL21 (DE3) for expression the recombined protein. The catalytic activity of the recombined protein was determined by Spectrophotometer and thin layer chromatography (TLC) methods., Result: The DNA sequence of PAL gene (designated as FdPAL, GenBank accession number: HM628904) was 2 583 bp in size, of which consisted two extrons and a single intron, and the full-length cDNA of FdPAL was 2 169 bp in size, which contained an ORF. The deduced protein of FdPAL contained 722 amino acids with calculated molecular weight (MW) of 78.31 kDa and an isoelectric point (pI) of 5.94. The SDS-PAGE results showed that the molecular weight of recombinant FdPAL protein was 75.37 kDa, which is consistent with the predictions. After 4 hours of induction, the enzymatic specific activity of FdPAL reached the summit, up to 4 386 nmol x g(-1) x min(-1). The reaction products were also identified by TLC, using L-Phe and trans-cinnamic acid as the internal standard., Conclusion: The PAL gene (both DNA sequence and full-length cDNA sequence) was cloned from F. dibotrys, and it has the same classic characters as other PALs in plants. The recombinant FdPAL was efficiently expressed in E. coli and had the activity for catalyzing the conversion from L-phenylalanine to cinnamic acid.