The rate of corneal graft rejection is still high for high-risk keratoplasty although immune suppression drug is routinely used.The role of traditional Chinese medicine in corneal transplantation is concerned gradually.Heijingpaichitang on the prevention and treatment of rats with high-risk corneal allograft rejection needs further study.Objective This study was to investigate the inhibitory effect of heijingpaichitang on high-risk corneal transplantation immune rejection in rats.Methods Sixteen female SD rats were used as the donors and 32female Wistar rats were served as recipients.The high-risk corneal trasplantation models were established by corneal suture in 32 Wistar rats,and then homogeneity variant SD-Wistar corneal transplantation was performed.The recipients were randomized into model control group,cyclosporinc A (CsA)group,heijingpaichitang group and CsA +heijingpaichitang group.CsA,heijingpaichitang and CsA + heijingpaichitang was orally administered 4 days after operation once per day for 15 days,and normal saline solution was used at the same way in the model control group.Ocular anterior segment reaction was examined under the slit lamp and corneal opacification,edema and neovasculation were scored based on Larkin' s criteria.Rejection index of the corneal graft was recorded and the graft survival time was calculated.The pathological examination of the corneal graft was carried out in all rats,and the inflammatory cells in the corneas and CD4+ cells in the periphery blood were assayed using flow cytometry.The use of the animals complied with ARVO Statement.Results Corneal graft rejection occurred in 10 days after operation in the model control group,12-13 days in the CsA group and heijingpaichitang group and 22 days in the CsA +heijingpaichitang group.Compared with model control group,the scores of the corneal opacification,corneal edema and neovascularization were significantly lower in the CsA group,heijingpaichitang group and CsA+heijingpaichitang group (P＜0.05),and all the scores were declined in the CsA+ heijingpaichitang group compared with CsA group and heijingpaichitang group(P＜0.01),but no significant differences were seen in the scores between the CsA group and heijingpaichitang group(P＞0.05).The mean survival time of grafts was (10.38 ±1.69)days in the model control group,(22.50 ± 3.07) days in the CsA + heijingpaichitang group,with the significant difference (t =-9.790,P =0.000).The pathological examination of graft showed that the lymphocytes and new blood vessels were less in the CsA+heijingpaichitang group compared with CsA group and heijingpaichitang group 15 days after operation.Flow cytometry verified that the number of lymphocytes in graft,CD4+cells and CD4+/CD8+ in periphery blood were significantly lower in the heijingpaichitang group,CsA group and CsA+heijingpaichitang group compared with model control group (P＜0.05).Conclusions Heijingpaichitang can inhibit immune rejection to certain extent in high-risk corneal transplantation rat and has a similar effect to 0.1％ CsA.Heijingpaichitang and 0.1％ CsA have a synergistic effect.

This study is to investigate the effect and mechanism of puerarin on DNA damage of HaCaT cells induced by UVB. Puerarin pre-treated cells were irradiated with UVB at 30 mJ x cm(-2). Twenty four hours after irradiation, DNA damage was detected by comet assay, ceramide was measured by thin layer chromatography and gas chromatography, intracellular free calcium ion was analyzed by flow cytometry, the phosphorylation level of p38 protein was examined by Western blotting method. Levels of DNA damage, ceramide, free calcium ion and p-p38 protein were elevated in UVB model cells. Contrary to the model group, all indicators above were reduced in all groups pre-treated by puerarin. Puerarin restrains the ceramide accumulation to block downstream p38 MAPK pathway and calcium ion rising, therefore reduces DNA damage in HaCaT cells induced by UVB.
Calcium ; metabolism ; Cell Line ; Ceramides ; metabolism ; DNA Damage ; drug effects ; radiation effects ; Down-Regulation ; Humans ; Isoflavones ; pharmacology ; Keratinocytes ; cytology ; metabolism ; Phosphorylation ; Signal Transduction ; drug effects ; Ultraviolet Rays ; adverse effects ; p38 Mitogen-Activated Protein Kinases ; metabolism

BACKGROUNDHedgehog (Hh) signaling plays an important role in both embryonic development and postnatal tissue homeostasis. Aberrant Hh activation results in a large variety of cancers. This study was designed to discover novel modulators in Hh signaling pathway.

METHODSWe performed yeast-two-hybrid screening and immunoprecipitation to identify the interaction of Nedd4 and Smo. To verify whether Nedd4 is involved in the regulation of Hh signaling, we monitored the activation of Gli-luciferase reporter by overexpressing Nedd4 together with Gli-luciferase reporter. In order to examine the role of endogenous Nedd4 in regulating Hh signaling, we used a short hairpin RNA (shRNA) interference strategy to silence the Nedd4 expression, and then perform dual-luciferase reporter assay. Statistical comparisons were performed by Student's t tests.

RESULTSWe showed that Nedd4 binds to Smo in the transfected HEK293 cells. Overexpression of Nedd4 alone did not significantly activate the Gli reporter compared to pcDNA3 control (Nedd4 group: dimethyl sulfoxide (DMSO), relative luciferase unit (RLU) 1.87 ± 0.41). However, Smo agonist (SAG)-stimulated activation of Gli-luciferase reporter was markedly potentiated in Nedd4 transfected cells (Nedd4 group: SAG, RLU 13.49 ± 1.04, P < 0.05), indicating that overexpression of Nedd4 increases Gli luciferase reporter activity and Nedd4-induced activation of Hh signaling is activity dependent. In Nedd4 knockdown NIH 3T3 cells, the luciferase reporter activity was measured basally and after SAG treatment. In scrambled cells, compared to DMSO, SAG could activate reporter activity by (4.16 ± 0.84)-fold. In Nedd4 knockdown cells, the luciferase reporter activation by SAG was significantly inhibited (SAG, RLU 1.72 ± 0.24, P < 0.05); knockdown of Nedd4 did not change the basal activity of luciferase activity (DMSO, RLU 0.86 ± 0.11), suggesting that the loss of Nedd4 expression diminishes Gli-dependent activity in the Hh pathway and the regulation of Nedd4 in the Hh signaling pathway is activity-dependent.

CONCLUSIONNedd4 positively regulates the Hh pathway and provides a potential target for inhibiting Hh signaling in cancer therapy.

Animals ; Endosomal Sorting Complexes Required for Transport ; physiology ; HEK293 Cells ; Hedgehog Proteins ; physiology ; Humans ; Mice ; NIH 3T3 Cells ; Nedd4 Ubiquitin Protein Ligases ; Receptors, G-Protein-Coupled ; physiology ; Signal Transduction ; physiology ; Smoothened Receptor ; Transcription Factors ; physiology ; Ubiquitin-Protein Ligases ; physiology ; Zinc Finger Protein GLI1

OBJECTIVETo investigate the effect of the RNAi and the chemotherapy drugs nolatrxed on the expression of thymidylate synthase(TS) and the growth of the colorectal carcinoma LOVO cells.

METHODSThe siRNA was constructed targeting the human TS gene, and then transfected into the human colorectal cancer LOVO cells. RT-PCR and Western blot technique were used to observe the TS gene and protein expression levels, and MTT was used to detect cell proliferation after silencing the TS gene. In addition, siRNA and nolatrxed were applied to the LOVO cells to observe the TS protein expression and cell growth.

RESULTSTS siRNA significantly reduced the expression of TS gene and protein in LOVO cells, and inhibited cell growth. The IC50 value of LOVO cells was (1.46±0.25) μmol/L in TS siRNA combined with nolatrexed group, (6.81±0.31) μmol/L in the negative control group, and (6.47±0.43) μmol/L in the single nolatrexed group. After treatment of TS siRNA combined with nolatrexed on LOVO cells for 36 hours, the apoptosis index was higher than that in single TS siRNA and nolatrexed[(62.12±0.89)% vs.(21.56±0.67)% and(40.51±0.83)%, both P<0.05].

CONCLUSIONTS siRNA can partly suppress the expression of TS gene in LOVO cells, inhibit cell proliferation, promote cell apoptosis and enhance cell sensitivity to apoptosis induced by nolatrexd.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Colorectal Neoplasms ; enzymology ; pathology ; Humans ; Quinazolines ; pharmacology ; RNA Interference ; Thymidylate Synthase ; genetics ; metabolism

Objective To investigate the effects of advanced glycation end-products(AGEPs)on the function of normal keratinocytes in vitro so as to explore the role of AGEPs in impaired wound healing. Methods Normal rat keratinocytes were incubated with different concentrations of AGEPs.After 48 hours of culturing,the cell proliferation rates were measured by MTT colorimetric determination.The cell cycle distributions and apoptosis were analyzed with flow cytometry,and the migration was investigated by 24-well fluorimetric cell migration assay kit by exposing to 100?g/ml AGEPs.Nuclear extracts from these cells were examined for binding of nucleotides containing NF-?B consensus by immunocytochemistry and EMSA in vitro.Results The proliferations of normal keratinocytes were significantly arrested and many cells were induced to early apoptosis compared with control ones(P＜0.05)by exposing to AGEPs for 48 hours. Meanwhile AGEPs also irritated keratinocytes migration compared with control ones(P＜0.05).Inhibiting the activation of NF-?B could partly recover the proliferation of keratinocytes,reverse apoptosis and attenu- ate migration.Conclusion AGEPs are correlated with the migration,proliferation and apoptosis of kera- tinocytes by NF-?B.

OBJECTIVETo investigate the predictive value of the ambulatory blood pressure monitoring parameters on left ventricular hypertrophy (LVH) and carotid artery intima-media thickness (IMT) in the hypertensives.

METHODSWe evaluated 147 hypertensive patients who were never treated regularly before. All patients underwent ultrasound examinations of the heart and the IMT of carotid arteries. We classified them as LVH group (n = 45) or no LVH group (n = 102), and as IMT increased group (n = 52) or no IMT increased group (n = 95). The record of medical history, physical examination and 24 h ambulatory blood pressure monitoring (ABPM) were performed in all the patients. The biochemical parameters such as blood lipids, glucose and so on were tested. Then the data comparison was made.

RESULTS(1) There were no significant differences in clinical manifestations and biochemical parameters between the LVH and no LVH groups (P > 0.05). Age (68.3 +/- 6.2) year vs (65.6 +/- 5.8) year, male 75.6% vs 66.7%, body mass index (24.1 +/- 4.1) vs (23.8 +/- 4.7) (kg/m(2)), diabetes mellitus and(or) impaired glucose tolerance 40.0% vs 38.2%, angina pectoris 42.3% vs 38.9%, cerebral vascular diseases 19.2% vs 15.7%, total cholesterol (5.40 +/- 1.42) vs (5.28 +/- 1.46) mmol/L, triglycerides (1.80 +/- 1.02) vs (1.74 +/- 1.08) mmol/L, low-density lipoprotein cholesterol (4.03 +/- 1.43) vs (4.06 +/- 1.48) mmol/L, high-density lipoprotein cholesterol (1.00 +/- 0.30) vs (0.99 +/- 0.26) mmol/L. (2) The parameters of ABPM in LVH group were higher than those in no LVH group. There were significant differences (P < 0.05) in 24 h mean systolic blood pressure (140.7 +/- 14.1) vs (128.3 +/- 12.3) mm Hg, 24 h mean diastolic blood pressure (86.4 +/- 8.9) vs (81.6 +/- 9.3) mm Hg, daytime mean systolic blood pressure (142.8 +/- 13.9) vs (130.9 +/- 11.1) mm Hg, daytime mean diastolic blood pressure (86.9 +/- 8.8) vs (83.4 +/- 9.0) mm Hg, nighttime mean systolic blood pressure (129.0 +/- 13.2) vs (114.6 +/- 11.4) mm Hg, nighttime mean diastolic blood pressure (77.2 +/- 9.4) vs (67.5 +/- 8.1) mm Hg, 24 h pulse pressure (54.2 +/- 10.2) vs (46.9 +/- 9.6) mm Hg, daytime pulse pressure (55.9 +/- 10.5) vs (47.5 +/- 9.1) mm Hg, nighttime pulse pressure (51.8 +/- 10.7) vs (47.1 +/- 8.7) mm Hg, 24 h systolic blood pressure variance (8.4 +/- 2.0) vs (7.2 +/- 1.9), 24 h diastolic blood pressure variance (9.5 +/- 2.2) vs (8.0 +/- 2.1), the non-dipper rhythm of ambulatory blood pressure 55.6% vs 25.5%. (3) There were also no significant differences in clinical manifestations between the IMT increased and no IMT increased group (P > 0.05). While there were significant differences between the IMT increased and no IMT increased group in those parameters of ABPM (P < 0.05).

CONCLUSIONThere were more LVH or IMT increased persons in the hypertensives whose ABPM parameters were abnormal.

Aged ; Aged, 80 and over ; Blood Pressure ; Blood Pressure Monitoring, Ambulatory ; Carotid Arteries ; diagnostic imaging ; pathology ; Female ; Humans ; Hypertension ; diagnostic imaging ; pathology ; physiopathology ; Hypertrophy, Left Ventricular ; diagnostic imaging ; pathology ; Male ; Middle Aged ; Predictive Value of Tests ; Ultrasonography

BACKGROUND: Compared with conventional total knee arthroplasty (TKA), for patients with genu varum or genu valgus, indications, prosthesis choice, surgical accuracy and perioperative management during TKA should be paid more attention. OBJECTIVE: To compare the clinical effectiveness of iASSIST-assisted TKA and traditional TKA in the treatment of genu varum or genu valgus. METHODS: Twenty-one patients with genu varum or genu valgus undergoing TKA were selected, and were then randomized into two groups: iASSIST-assisted TKA (group A) or traditional TKA (group B). The surgical accuracy was compared between two groups by measuring the knee valgus angle of patients, and the angle between prosthesis components on the coronal and sagittal planes. Besides, the intraoperative blood loss, operation time, postoperative drainage, restored alignment, postoperative complications, hospitalization time, as well as Visual Analogue Scale and American Knee Society scores were recorded to assess the clinical effectiveness. RESULTS AND CONCLUSION: (1) The deviation of hip-knee-ankle angle, frontal femoral component angle, frontal tibial component angle, lateral femoral component angle and lateral femoral component angle in the two groups was less than 3°, and the group A exhibited better corrective efficacy. (2) The operation time in the group A was significantly longer than that in the group B; the hospitalization time in the group was significantly shorter than that in the group B; the restored alignment in the group A was significantly superior to that in the group B;the postoperative drainage, as well as Visual Analogue Scale and American Knee Society scores at 1 day and 1 week postoperatively in the group B were significantly superior to those in the group A (all P < 0.05). There were no significant differences in the intraoperative blood loss, incidence of complications, as well as Visual Analogue Scale and American Knee Society scores at postoperative 2 weeks and 1 and 3 months between two groups (P > 0.05). (3) These results manifest that for the patients with genu varum or genu valgus, both iASSIST-assisted TKA and traditional TKA can obtain satisfactory surgical accuracy, and the former has advantage in restricting the alignment of lower limbs. However, the long-term efficacy needs to be explored in depth.

Objective To investigate the technological feasibility,efficacy and morbidity of cr guided ~(125)Ⅰseed implantation for recurrent rectum cancer.Methods Twenty-three patients with recurrent rectum cancer were treated with cr guided interstitial ~(125)Ⅰseed brachytherapy.In 20 patients the procedure was performed under epidural anesthesia and 3 patients under local anesthesia.Treatment planning system was used to calculate the number of seeds,the space distribution and the introduction of the seeding needles.Matched peripheral dose (MPD) of ~(125)Ⅰseed implantation ranged from 90-120 Gy for patients who had had external radiotherapy,and 140- 160Gy for those who had not.The planning target volume(PTV)was clinical target volume(CTV)plus 1 cm margin.The range of radioactivity of the ~(125)Ⅰseeds was 18.5-25.9 MBq.All these 23 patients had CT scan at 5 mm intervals after implantation for quality evaluation,together with routine chest,pelvic X-ray films within 24-48 hours after seed implantation.Three patients received three-dimensioual conformal radiation therapy(3DCRT) to a total dose of45-50 Gy,with 2-3 Gy/f.Follow-up time was from 3 to 28 months.Results All patients was able to tolerate seed implantation well.Complete pain relief was observed in 12/15,and partial relief in 2/15 and no response in 1/15,with a response rate of 93%.The local control rate was 87%.The 1-and 2-year survival rate was 93% and 50% respectively.Two of four patients have died of dissemination to the lung after 8 and 12 months. One seed has migrated into the pelvis without causing any untoward morbidity.Conclusion CT guided ~(125)Ⅰseed implantation for recurrent rectum cancer is safe,minimally invasive,causing only mild morbidity.It possesses a high efficacy,yet it should be given in combination with external beam radiation and chemotherapy,should distant metastasis be observed.

BACKGROUNDCortical spreading depression can cause migraine attack, and up-regulate matrix metalloproteinase-9 (MMP-9) expression in animal. This study aimed to determine the impact on the structure and function of the blood-brain barrier by measuring plasma MMP-9 levels in patients at the acute and late stages of migraine attacks in order to elucidate the pathological mechanisms involved.

METHODSWe recruited a case-control cohort of 38 adult migraine patients and 20 age- and gender-matched healthy control subjects. Five milliliter blood samples were collected at the acute and late stages of migraine (days 1 - 7), and also from the control subjects. Solid phase double antibody sandwich enzyme-linked immunosorbent assay was used to determine plasma MMP-9 levels. Statistical analysis was performed using the SAS version 9.1.

RESULTSInitial plasma MMP-9 levels of migraine patients were significantly higher than those of controls ((12.612 ± 0.016) µg/L vs. (6.069 ± 0.023) µg/L, respectively, P < 0.05). High MMP-9 expression was observed during days 1 - 6 of migraine attacks, with highest expression occurring on day 3 ((17.524 ± 0.035) µg/L). During attacks, MMP-9 levels were similar in migraine patients with and without aura (P > 0.05); in addition, levels were not correlated with degree of headache pain (P > 0.05).

CONCLUSIONSWe hypothesize that migraine could lead to increased plasma MMP-9 levels resulting in blood-brain barrier damage. MMP-9 levels increase during days 1 - 6 of migraine attacks, peaking on day 3. Therefore, MMP-9 could be used as a biological marker to guide treatment of migraine attacks.

Adult ; Blood-Brain Barrier ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Male ; Matrix Metalloproteinase 9 ; blood ; Middle Aged ; Migraine Disorders ; physiopathology ; Young Adult

Thyroid cancer is the one of the most common endocrine tumors. The biological behaviors and prognoses of the thyroid cancer of different histological types remarkably differ. The highly invasive thyroid cancer responds poorly to traditional therapies. Recent research advances in the molecular mechanisms of the pathogenesis of thyroid cancer have revealed the roles of many genetic and epigenetic variations such as gene mutation, abnormal gene amplification, and abnormal gene methylation in the development of thyroid cancer, which provides new insights in the molecular diagnosis, prognosis, and target therapy of the thyroid cancer.
Carcinoma ; genetics ; Humans ; Mutation ; Signal Transduction ; Thyroid Neoplasms ; genetics