Effect of doxycycline on the proliferation of bovine corneal myofibroblast in vitro / 中华实验眼科杂志

Background Doxycycline is a broad spectrum antibiotic,and it is frequently used in the treatment of ocular surface diseases.Objective The purpose of the present study was to investigate the effect of doxycycline on the inhibition of cell proliferation in bovine corneal myofibroblasts in vitro and assess its contribution to ocular surface repairing mechanism.Methods Six fresh bovine corneas were collected.The corneal stromal layer was isolated by two-step method of 1.0 g/L and 2.0 g/L collegenase-1.Isolated cells were plated at mantaryay culture flask in 10％ FBS of RPMI-1640.Vimentin and alpha-smooth muscle actin (α-SMA) organization were evaluated by immunocytochemistry,and the cells with influoresccence staining for vimentin and α-SMA were identified as the corneal myofibroblasts.Doxycycline at the concentrations of 10,20,40,60,80 mg/L was added to the medium,respectively,in different concentrations of doxycycline groups.Dexamethasone (120 mg/L)was used in the same way in the positive control group,and no drug was used in the negative control group.Cell proliferation was evaluated by MTT and the cell cycle was analyzed by BD FACScan flow cytometer assay 24 hours and 48 hours after addition of any drug.Results The cells grew well and showed the positive response for vimentin and α-SMA.MTT assay showed that the A570values of bovine corneal myofibroblasts were gradually declined with the increase of the concentration of doxycycline and lapse of active time,showing statistically significant difference (Fconcentration =1233.778,P＜0.001 ; Ftime =227.564,P ＜ 0.001).And the difference between the two factors was also statistically significant (Ftime*concentration =51.656,P＜0.001).Flow cytometry cell cycle analysis showed that 24 hours after 10,20,40,60,80 mg/L doxycycline treated,the perentage of of corneal myofibroblast cell in G0-G1 phase was 82.85％,84.36％,85.18％,87.12 ％ and 89.31％,showing significant increase in comparison with 63.89％ of the negative control group (all P＜0.05),and that of 40 mg/L doxycycline group was near the positive control group.Forty-eight hours after 10,20,40,60,80 mg/L doxycycline treated,the perentage of of corneal myofibroblast cell in G0-G1 phase was 82.78％,86.15％,88.23％,89.57％,93.00％,with significant increase in comparison with 70.17％ of the negative control group (all P ＜ 0.01),and that of 40 mg/L doxycycline group was near the positive control group.Conclusions The growth of the bovine corneal myofibroblasts is inhibited by doxycycline in time-and dosedependent manner in the range from 10 mg/L to 80 mg/L,and 40 mg/L of doxycycline has an obviously inhibitory action as 120 mg/L dexamethasone.

Effect of depside salt from salvia miltiorrhizae in repairing advanced glycation end products-induced late endothelial progenitor cell dysfunction and its molecular mechanism / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To investigate the effects of depside salt from salvia miltiorrhizae (DSSM) in repairing advanced glycation end products (AGE)-induced late endothelial progenitor cell (EPC) dysfunction, and its possible molecular mechanism.</p><p><b>METHODS</b>Mononuclear cells (MNCs) were separated using density gradient centrifugation from human umbilical cord blood, and cultured with EGM-2-MV culture fluid to late EPCs. Then the EPCs were divided into 5 groups: Group A incubated with 200 microg/mL AGE-modified bovine serum albumin (AGE-albumin) alone (A), Groups B, C and D with equal dosage of AGE-albumin plus DSSM at different dosages (0.1 microg/mL, 1 microg/mL, and 10 microg/mL), Group E with 200 microg/mL of unmodified-AGE. The late EPCs apoptosis was detected by Annexin V+/PI double-stain, angiogenic capacity was detected by ECMatrix-gel, mRNA expressions of the receptor for AGE (RAGE) and endothelial nitric oxide synthase (eNOS) were measured by reverse-transcriptase polymerase chain reaction (RT-PCR), and the protein expressions of RAGE, eNOS and protein kinase (Akt) were measured by Western blot.</p><p><b>RESULTS</b>Compared with Group E, in Group A, the Annexin V+/PI- ratio and expression of RAGE in EPCs increased, the angiogenic capacity, mRNA and protein expressions of eNOS, and protein expression of Akt decreased significantly. These abnormal changes in Groups C and D were significantly smaller than those in Group A (P < 0.05 or P < 0.01). And all the indices in Group D were adjacent to those in Group E, showing insignificant difference between the two groups (P > 0.05).</p><p><b>CONCLUSIONS</b>AGE could injure the function of EPCs, revealing increase of cell apoptosis and migration, deprivation of angiogenic capacity in vitro. DSSM could repair the EPCs dysfunction induced by AGE-albumin. Up-regulation of eNOS and Akt in these cells may be involved in the mechanism.</p>

Relationship between Chinese medicine syndrome type and psychological factor in patients with functional dyspepsia / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To explore the relationship between Chinese medicine syndrome type and psychological factor in patients with functional dyspepsia (FD).</p><p><b>METHODS</b>With an epidemiologic method adopted, 297 FD patients received psychologic mensuration and their Chinese medicine syndrome type was differentiated. The distribution of Chinese medicine syndrome type in FD patients was studied and the differences among various types were analyzed using self-rating depression scale (SDS) and self-rating anxiety scale (SAS).</p><p><b>RESULTS</b>(1) Patients' Chinese medicine syndrome could be differentiated into 5 types: the Pi-deficiency qi-stagnancy type (I), the Gan-qi accumulation type (II), the Gan-qi invading Wei type (III), the dampness-heat stagnating in Wei type (IV) and the Pi-Wei qi-deficiency type (V). Patients of type I (96 cases, 32.3%) held the dominant share. (2) Depressive and anxiety states presented in patients with various syndrome types, among them, patients of type II held the highest percentage of depressive status (30 cases, 62.5%), type III held the highest percentage of anxiety state (19 cases, 35.8%), while type IV possessed the lowest percentages of both. (3) Analysis between symptoms and syndrome types showed that post-prandial fullness presented in most patients of types V and I; early satiation presented more prominently in patients of type V; upper abdominal pain presented frequently in patients of types II and V, and upper abdominal burning sensation presented more evidently in patients of type IV. (4) Comparisons of SDS and SAS scores in patients with different syndrome types showed that the highest SDS score presented in type II, highest SAS score presented in type III; and the lowest scores of SDS and SAS all presented in type IV.</p><p><b>CONCLUSIONS</b>Psychological states are different in FD patients with various syndrome types. The Chinese medicine pathogenetic mechanisms of FD is complex in deficiency/excessive nature, and the condition of disease is closely related with organs Gan and Pi.</p>