OBJECTIVETo study the modulatory effect of insulin on apoptosis of skeletal myoblast (L6 cells) by serum of scalded rat and its mechanism.

METHODSL6 cells cultured with DMEM medium containing 10% FBS were divided into control (C, added with 20% normal rat serum), serum from rat with scald injury (S, added with 20% serum from scalded rat), insulin (I, added with 20% normal rat serum and 100 nmol/L insulin), and serum of scalded rat + insulin (SI, added with 20% serum of scalded rat + 100 nmol/L insulin) groups according to the random number table. After being cultured for 48 hours, apoptosis was observed with Hoechst 33258 staining and its number counted, annexin V -FITC/PI double-labeling method was used to assess apoptosis rate, the protein levels of phosphorylated (p-) Akt, p-PI3K, Bax, Bcl-2, and active caspase-3 were determined by Western blotting. Data were processed with grouped or paired t test.

RESULTS(1) The amount of apoptosis with typical morphological change in S group [(59.6 +/- 3.9) per visual field] was more than that in C, I, and SI groups [(4.9 +/- 2.6), (5.5 +/- 2.1), (19.7 +/- 2.3) per visual field, with t value respectively 28.53, 29.86, 21.53, P values all below 0.01]. (2) Apoptotic rate in S group was (18.5 +/- 1.8)%, which was markedly higher than that in C, I, and SI groups [(1.1 +/- 0.6)%, (1.5 +/- 0.3)%, (7.8 +/- 0.6)%, with t value respectively 22.41, 22.83, 13.92, P values all below 0.01]. (3) Compared with those in C group, the protein levels of Bax and active caspase-3 in S group were up-regulated (1.12 +/- 0.63 vs. 0.16 +/- 0.03, 2.15 +/- 0.51 vs. 0.21 +/- 0.03, with t value respectively 3.80, 10.69, P values all below 0.01), the protein level of p-Akt was lowered (0.20 +/- 0.03 vs. 0.42 +/- 0.07, t = -8.46, P < 0.01), and the protein levels of p-PI3K and Bcl-2 showed no statistical difference (0.19 +/- 0.03 vs. 0.26 +/- 0.09, 0.17 +/- 0.03 vs. 0.28 +/- 0.07, with t value respectively -2.73, - 1.14, P values all above 0.05). The protein levels of Bax (0.40 +/- 0.14) and active caspase-3 (0.83 +/- 0.18) in SI group were lowered (t = -3.23, P < 0.05; t = 6.66, P < 0.01) and the protein levels of p-Akt, Bcl-2, and p-PI3K in SI group were elevated (0.39 +/- 0.10, 0.78 +/- 0.03, 0.47 +/- 0.12, with t value respectively 4.07, 18.71, 5.05, P < 0.05 or P < 0.01) as compared with those in S group.

CONCLUSIONSSerum from scalded rat can induce apoptosis in skeletal myoblast, and the effect can be inhibited by insulin through PI3K/Akt signal pathway.

Animals ; Apoptosis ; drug effects ; Burns ; blood ; metabolism ; pathology ; Caspase 3 ; metabolism ; Cell Line ; Insulin ; pharmacology ; Male ; Myoblasts, Skeletal ; cytology ; drug effects ; pathology ; Rats ; Rats, Wistar ; Serum ; immunology ; Signal Transduction ; bcl-2-Associated X Protein ; metabolism

AIM: To investigate the effect of tumor necrosis factor receptor associated factor 6 (TRAF6) on proliferation, apoptosis and invasion of retinoblastoma Y79 cells. METHODS: The Y79 cells were divided into three groups:blank control group, negative control group and TRAF6 siRNA group. After TRAF6 siRNA transfection, the levels of TRAF6 mRNA and protein in Y79 cells were examined by RT-PCR and Western blotting. MTT assay was used to detect cell proliferation. Flow cytometry was employed to detect changes in cell cycle and apoptosis. Cell invasiveness was detected by the Transwell method. RESULTS: Expression of TRAF6 mRNA and protein in the TRAF6 siRNA group significantly decreased compared with the negative and blank control groups. Following the silencing of TRAF6,cell proliferation was inhibited and the apoptosis rate increased; the cell cycle was arrested at G0/G1 phase; the number of cells in S phase was reduced, while the invasion ability of cancer cells decreased. CONCLUSION: Silencing TRAF6 may inhibit the proliferation of Y79 cells, promote cell apoptosis, arrest the cell cycle at G0/G1 phase and decrease the invasive ability. Thus,TRAF6 may be a potential target in therapy for retinoblastoma.

Background Accurate evaluation of response following chemotherapy treatment is essential for surgical decision making in patients with breast cancer.Modalities that have been used to monitor response to neo-adjuvant chemotherapy (NAC) include physical examination (PE),ultrasound (US),and magnetic resonance imaging (MRI).The purpose of this study was to evaluate the accuracy of PE,US,and MRI in predicting the response to NAC in patients with breast cancer.Methods According to the response evaluation criteria in solid tumors guidelines,the largest unidimensional measurement of the tumor diameter evaluated by PE,US,and MRI before and after NAC was classified into four grades,including clinical complete response,clinical partial response,clinical progressive disease,clinical stable disease,and compared with the final histopathological examination.Results Of the 64 patients who received NAC,the pathologic complete response (pCR) was shown in 13 of 64 patients (20％).The sensitivity of PE,US,and MRI in predicting the major pathologic response was 73％,75％,and 80％,respectively,and the specificity was 45％,50％,and 50％ respectively.For predicting a pCR,the sensitivity of PE,US,and MRI was 46％,46％,and 39％,respectively,and the specificity was 65％,98％,and 92％ respectively.Conclusions Compared with final pathologic findings,all these three clinical and imaging modalities tended to obviously underestimate the pCR rate.A more appropriate,universal,and practical standard by clinical and imaging modalities in predicting the response to neo-adjuvant chemotherapy in vivo is essential.

Phytochemical investigation on the stems of Ilex asprella by using various chromatographic techniques led to the isolation of 13 compounds. By spectroscopic analyses and comparisons the spectral data with those in literatures, these compounds were identified as salicifoneoliganol(1), rel-(7R,8S)-3,3',5-trimethoxy-4',7-epoxy-8,5'-neolignan-4,9,9'-triol 9-β-D-glucopyranoside(2),(+)-cycloolivil(3),(+)-syringaresinol-4'-O-β-D-monoglucoside(4), liriodendrin(5), caffeic acid (6), 3,4-dihydroxy-5-methoxybenzaldehyde(7), benzene-1,2,4-triol(8), 3,4,5-trimethoxyphenyl-1-O-β-D-apiofuranosyl(1″→6')-glucopyranoside(9), aeculetin(10), cryptochlorogenic acid ethyl ester(11), chlorogenic acid ethyl ester(12), and rel-5-(3S,8S-dihydroxy-1R,5S-dimethyl-7-oxa-6-oxobicyclo [3,2,1]oct-8-yl)-3-methyl-2Z,4E-pentadienoic acid(13). Among them, compounds 7, 8, 11, and 13 were isolated from genus Ilex for the first time, and 1-3, 9, 10, and 12 were isolated from this speciesfor the first time. The anti-inflammatory assay results of these compounds showed that compounds 1 and 9 showed moderate inhibitory effect against NO production in RAW 267. 4 cells with IC₅₀ values of 35.7 and 50.6 μmol•L⁻¹, in vitro respectively, whereas compound 10 showed weak inhibition(IC₅₀ value 98.7 μmol•L⁻¹).

OBJECTIVE: To study the mechanism of Shengmai Injection (SMI, ) on anti-sepsis and protective activities of intestinal mucosal barrier. METHODS: The contents of 11 active components of SMI including ginsenoside Rb1, Rb2, Rb3, Rd, Re, Rf, Rg1, Rg2, ophioposide D, schisandrol A and schisantherin A were determined using ultra-performance liquid chromatography. Fifty mice were randomly divided into the blank, the model, the low-, medium- and high-dose SMI groups (0.375, 0.75, 1.5 mL/kg, respectively) by random number table, 10 mice in each group. In SMI group, SMI was administrated to mice daily via tail vein injection for 3 consecutive days, while the mice in the blank and model groups were given 0.1 mL of normal saline. One hour after the last SMI administration, except the blank group, the mice in other groups were intraperitoneally injected with lipopolysaccharide (LPS) saline solution (2 mL/kg) at a dosage of 5 mL/kg for development of endotoxemia mice model. The mice in the blank group were given the same volume of normal saline. Inflammatory factors including interferon-γ (INF-γ), tumor necrosis factor-α (TNF-α), interleukin (IL)-2 and IL-10 were measured by flow cytometry. Myosin light-chain kinase (MLCK), nuclear factor κB (NF-κB) levels, and change of Occludin proteins in jejunum samples were analyzed by Western blot. RESULTS: The decreasing trends of INF-γ, TNF-α and IL-2 were found in serum of SMI treatment groups. In SMI-treated mice, the content of Occludin increased and MLCK protein decreased compared with the model group (P<0.05 or P<0.01). The content of cellular and nuclear NF-κB did not change significantly (P>0.05). CONCLUSION SMI may exert its anti-sepsis activity mainly through NF-κB-pro-inflammatory factor-MLCK-TJ cascade.
Animals ; Drug Combinations ; Drugs, Chinese Herbal ; Mice ; NF-kappa B/metabolism* ; Occludin ; Saline Solution ; Sepsis/drug therapy* ; Tumor Necrosis Factor-alpha/metabolism*

In recent years, the real-world studies (RWS) have attracted extensive attention, and the real-world evidence (RWE) has been accepted to support the drug development in China and abroad. However, there is still a lack of standards for the evaluation of the quality of RWE. It is necessary to formulate a quality evaluation and reporting specification for RWE especially in traditional Chinese medicine (TCM). To this end, under the guidance of China Association of Chinese Medicine, the Quality Evaluation and Reporting Specification for Real-World Evidence of Traditional Chinese Medicine (QUERST) Group, including 24 experts (clinical epidemiologists, clinicians, pharmacologists, ethical reviewer and statisticians), was established to develop the specification. This specification contains the listing of classification of RWS design and RWE, the general principles and methods of RWE quality evaluation (26 tools or scales), 25 types of bias in RWS, the special considerations in evaluating the quality of RWE of TCM, and the 19 reporting standards of RWE. This specification aims to propose the quality evaluation principles and key points of RWE, and provide guidance for the proper use of RWE in the development of TCM new drugs.
Medicine, Chinese Traditional ; China