OBJECTIVETo investigate the effect of advanced glycosylation end products (AGE) on cell cycle of epidermal keratinocyte and its possible signal pathway.

METHODS150 mg/L AGE-human serum albumin (AGE-HSA) was prepared in vitro. Primary cultured keratinocytes in logarithmic growth phase were harvested and divided randomly into: A group [with treatment of defined keratinocyte-SFM (DK-SFM) serum-free medium], B group (with treatment of DK-SFM medium including 150 mg/L AGE-HSA), C group (with DK-SFM medium after treatment of U0126) and group D (with D K-SFM medium including 150 mg/L AGE-HSA after treatment of U0126). Cell cycle distributions were analyzed by flow cytometer. The protein levels of cyclin D1, cyclin B1, CDK4 and p44/42 MAPK were measured by Western blot.

RESULTSCompared with those of A group, the percentage of S-phase and G2/M-phase keratinocytes were decreased obviously in B group, the percentages of G2/M -phase keratinocytes showed the same tendency in C and D groups [(9.7 +/- 1.1)% , (9.8 +/- 0.7)%, respectively, P <0.05]. Compared with that of A group, the expression of cyclin D1 were decreased significantly in other groups, among which a weak expression was showed in D group. There was no obvious difference between A and B groups in CDK4, or cyclin B1 and p44/42 MAPK protein levels ,which were significantly higher than those in C and D groups.

CONCLUSIONAGEs inhibit the progress of cell cycle of keratinocytes by downregulation of cyclin D1 expression.

Animals ; Cell Cycle ; Cyclin D1 ; metabolism ; Epidermis ; cytology ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Glycation End Products, Advanced ; metabolism ; pharmacology ; Keratinocytes ; cytology ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Signal Transduction

OBJECTIVETo analyze the prevalence, clinical features, diagnosis, potential risk factors, anti-tuberculosis treatment efficacy and prognosis of the patients with leukemia complicated with active tuberculosis (TB).

METHODSA retrospective study was performed to analyze the clinical characteristics, relevant examination data, diagnosis methods and follow-up data about 44 leukemia cases complicated with active TB from January 2006 to December 2011 in our single center.

RESULTSThe prevalence of leukemia complicated with active TB was 1.70% (pulmonary TB 1.35%, extra-pulmonary TB 0.35%) and no statistically significant difference was found between each subgroup of acute and chronic leukemia groups (P>0.05). Most of the patients were men, with a male to female ratio of 2.14:1, the median age of 40 years old (range 16 to 78), presenting as atypical clinical manifestations, such as high fever, cough, and so on. Eighteen patients (40.9%) were diagnosed with definitely etiological evidence while the other 26 patients (59.1%) were diagnosed clinically. The extra-pulmonary TB group had a higher purified protein derivative (PPD) test positive rate than that of the pulmonary TB group (88.9% vs 42.9%, P=0.020). The chest CT and T-cell spot of tuberculosis test (T-SPOT.TB) were helpful tools for diagnosis. The potential risk factors included age, sex, nutritional status, neutropenia, decreased cellular immunity, type and course of leukemia, etc. The significant differences in age, gender, administration route of immunosuppressive drugs were found between neutropenic and non-neutropenic groups (P<0.05). The efficacy of first-line anti-tuberculosis therapy was 83.7% and the total course to cure TB was around 12 months. Four patients were dead due to pulmonary TB with a 9.1% attributable mortality.

CONCLUSIONThe prevalence of leukemia complicated with active TB is higher than the general population in our single center. The main characteristics including various potential risk factors, atypical clinical features, diagnoses mainly made by clinical features were found in our patients with leukemia complicated with active TB. However, it showed that these patients demonstrated good responses to the first-line anti-tuberculosis therapy and relative lower attributable mortality.

Adolescent ; Adult ; Aged ; Female ; Humans ; Leukemia ; complications ; epidemiology ; Male ; Middle Aged ; Prevalence ; Retrospective Studies ; Risk Factors ; Tuberculosis ; drug therapy ; epidemiology ; Young Adult

Impaired wound healing in diabetes is a significant clinical problem which is thought to be associated with neuropathy and angiopathy previously . The present study indicates that accumulation of glucose and glycometabolic products in skin tissue, as the result of glycometabolic disorders, which contributes to cutaneous environmental alterations in diabetes mellitus, and subsequently induces the abnormal cell behaviors, cytokine alteration and matrix modification. Thus, diabetic neuropathy and angiopathy might be regarded as the pathological outcome of cutaneous environmental alterations. In conclusion, glycometabolism disorders could be described as one of the initial events for the alteration involving in the underlying cutaneous disorder which impair healing process. The related research focuses on the initial event of controlling disorders in wound healing and therefore contribute to providing the strategy of treatment as based on these approaches.
Diabetes Mellitus ; metabolism ; physiopathology ; Humans ; Skin ; metabolism ; Wound Healing

OBJECTIVETo investigate the rule and possible mechanism of epidermal proliferation in wound edge of deep partial thickness scald injury in rat.

METHODSTwenty-four Sprague-Dawley rats inflicted with deep partial thickness scald were randomized into pre-scalding, 3 post-scalding day (PSD), 7PSD and 14PSD groups, with 6 rats in each group. Skin specimens from the wound edge were harvested for the observation of the histological characteristics of the epidermis. Cell cycles of epidermal cells were analyzed with flow cytometry. The expressions of cyclin D1, cyclin B1, cdk4 and the histone H1 kinase activity of MPF in epidermal cells were determined by Western blotting.

RESULTSAugmentation of nuclei and nucleoli was found in the epidermal cells from the wound edge in 3PSD group, while increased number of epidermal cells with obviously augmented nuclei and nucleoli were found in 14PSD group. The percentage of the cells in S phase increased in 14 PSD group. The percentage of epidermal cells in G2/M phase began to increase in 3PSD group, and that in 7PSD (4.5 +/- 0.6) and 14PSD (5.4 +/- 1.0) groups were obviously higher than that in pre-scalding group (2.9 +/- 1.1, P < 0.05). The expression of cyclin D1 increased significantly in 3PSD group. The expression of cdk4 decreased in 3PSD group, but began to increase in 14PSD group. There was no difference in the expression of cyclin B1 among groups. The MPF activity was significantly increased in 14PSD group.

CONCLUSIONThere was enhanced DNA synthesis and mitosis in epidermal cells of rats with deep partial scald during early post-scald stage, and active proliferation of epidermal cells was observed on 14PSD. The expression of cyclinD1/cdk4 complex and the activity of MPF increased since 14PSD, indicating that there was a special regulative pattern during wound healing.

Animals ; Burns ; pathology ; Cell Cycle ; Cell Proliferation ; Disease Models, Animal ; Epithelial Cells ; cytology ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Soft Tissue Injuries ; pathology ; Wound Healing

OBJECTIVETo investigate the possible mechanism of L-arginine supplementation on the angiogenesis of burn wounds in diabetic rats.

METHODSOne hundred male Sprague-Dawley (SD) rats were used in the study and were randomly divided into A (scalding control, n = 25), B (scalding of the rats with diabetes, n = 25), C (L-glycine control, n = 25) and D (L-arginine supplementation, n = 25) groups. Diabetes was produced by intra-peritoneal injection of streptozotocin (STZ) in B, C and D groups. The rats in C and D groups were gavaged with L-glycine and L-arginine in dose of 200 mg.kg(-1).d(-1), respectively. The glucose content of the back skin tissue was determined for five rats in each group eight weeks after STZ administration. Deep partial thickness scalding of 20% TBSA was engendered on the back in the other 80 rats. The wound area, wound healing rate, and microvascular density with CD34 immunohistochemistry staining were determined on 3rd, 7th, 14th, and 21st post scalding days (PSDs), In addition, the amount of nitric oxide (NO) released from the wound tissue and the tissue contents of vascular endothelial growth factor (VEGF) and transforming growth factor beta1 (TGF-beta1) from wound were determined at the above time points.

RESULTSCompared to those in group B, the wound healing rate in group D increased significantly since the 7th PSD [(44.10 +/- 3.50)%, P < 0.05], and the wound MVD value was increased significantly at all postburn time points. Furthermore, the levels of VEGF, NO and TGF-beta1 in the wound tissue was also increased significantly, while the glucose content in the cutaneous tissue was decreased to (1.380 +/- 0.120) mg/g.

CONCLUSIONL-arginine supplementation could be beneficial to the angiogenesis in the burn wound of the rats with diabetes, as well as to wound healing by increasing the synthesis and the release of VEGF, NO and TGF-beta1 from burn wound and by decreasing the glucose content in the cutaneous tissue of diabetic rats.

Animals ; Arginine ; therapeutic use ; Blood Glucose ; metabolism ; Burns ; metabolism ; therapy ; Diabetes Mellitus, Experimental ; metabolism ; therapy ; Male ; Neovascularization, Physiologic ; drug effects ; Nitric Oxide ; metabolism ; Rats ; Rats, Sprague-Dawley ; Transforming Growth Factor beta1 ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism ; Wound Healing ; physiology

OBJECTIVETo understand the influence of accumulation of advanced glycosylation end products (AGE) on wound healing of burn rats complicated with diabetes.

METHODSSeventy-five SD rats were divided into control, diabetes, and aminoguanidine-interfered groups in completely randomized method, with 25 rats in each group. All rats were subjected to deep partial-thickness scald. Diabetes was reproduced in rats of diabetes and aminoguanidine-interfered groups. Rats in aminoguanidine-interfered group were fed with 100 mg x kg(-1) xd (-1) aminoguanidine. Rats were sacrificed on post-scald day (PSD) 0, 3, 7, 14, and 21, and portrait of the wounds were taken. Full-thickness skin tissue specimens were obtained for determination. Specimens of epidermis from back of SD rats were obtained for KC cultivation and verification. Wound healing rate, glucose content in skin tissue, morphologic change in wound tissue, AGE distribution in skin tissue, influence of AGE on proliferation and apoptosis of KC were observed.

RESULTSWound healing rate of rats was respectively lower in diabetes group than that in control group on PSD 7, 14, and 21 (P < 0.01), but it was obviously higher in aminoguanidine-interfered group than that in the former 2 groups (P < 0.01). Glucose content of rat skin in diabetes group was (2.62 +/- 0.19) mmol/g, and it was (2.58 +/- 0.07) mmol/g in aminoguanidine-interfered group, both higher than that in control group [(1.04 +/- 0.09) mmol/g, P < 0.01]. In control group, limited intensive infiltration of inflammatory cells was found in the wound with necrotic tissue formation which fell off in time, and with no obvious delay of wound healing. In diabetes group, infiltration of inflammatory cells in wounds of rats appeared slowly, but diffusely and persistently; necrotic tissue formed and fell off late in time, with obvious delay of wound healing. In aminoguanidine-interfered group, intensive infiltration of inflammatory cells was observed in time, and the time of necrotic tissue formation and sloughing, and wound healing were respectively earlier than that in diabetes group. Sporadic disposition of small amount of AGE was found in rats in control group. AGE accumulation increased significantly in rats in diabetes group. AGE content decreased significantly in rats in aminoguanidine-interfered group after administration of aminoguanidine. KC proliferation decreased significantly in concentration dependent manner 48 hours after AGE stimulation. Absorbance value of AGE decreased in each AGE-interfered group (P < 0.01). Early Annexin-V positive apoptotic KC rate was obviously higher in 100 ug/mL AGE-interfered group (15.1 +/- 2.3)% than that in control group [(11.2 +/- 1.2)%, P < 0.05]. There was no statistical significance between 100 ug/mL AGE-interfered group (14.3 +/- 3.5)% and control group (15.2 +/- 2.4)% in respect of the rate of double-positive cells apoptosis at final stage (P > 0.05).

CONCLUSIONSHyperglycemia may inhibit proliferation of repairing cells such as KC through AGE accumulation, thus impedes wound healing. Reduction of AGE accumulation could ameliorate wound healing delay due to diabetes.

Animals ; Blood Glucose ; metabolism ; Burns ; complications ; metabolism ; Diabetes Mellitus, Experimental ; complications ; metabolism ; Glycation End Products, Advanced ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Wound Healing

OBJECTIVETo analyze the relationship between cutaneous glycometabolic disorders and cutaneous neuropathy in diabetic rats, and to look for the mechanism of neuropathy and impaired wound healing.

METHODSEighty male SD rats were randomly divided into the normal control group (NC, n = 20), diabetic group (D, n = 20), aminoguanidine-interfered group (AI, n = 20), and insulin-interfered group (II, n = 20) by drawing lots. Diabetes was reproduced in rats of D, AI, and II groups with intraperitoneal injection of streptozotocin (STZ). Then, rats in AI group were fed with 100 mg×kg(-1)×d(-1) aminoguanidine, while rats in II group were subcutaneously injected with insulin for satisfactory control of blood glucose. Changes in mechanical and heat pain thresholds of pad of hind limb were measured at post injection week (PIW) 2, 4, 8. Skin specimens were collected during PIW 2-8 from pads for determination of contents of glucose, advanced glycation end product (AGE), substance P (SP), calcitonin gene-related peptide (CGRP), and observation of distribution and ultrastructure of skin nerve fibers. Data were processed with t test.

RESULTSThe mechanical and heat pain thresholds in D group at PIW 2 [(6.3 ± 1.5) g, (6.0 ± 0.9) s, respectively ] were obviously lower than those in NC group [(13.0 ± 3.2) g, (10.3 ± 1.2) s, with t value respectively 2.71, 3.42, P values all below 0.05]. Contents of glucose and AGE in skin tissue in D group were significantly increased when compared with those in NC group, especially at PIW 8 [(2.85 ± 0.33) mg/g, (31.7 ± 3.2) U/mg of hydroxyproline vs. (0.82 ± 0.22) mg/g, (22.2 ± 1.9) U/mg of hydroxyproline, with t value respectively 1.65, 6.47, P values all below 0.01]. The myelinated nerve fibers were edematous and degenerated, with axons compressed, while the unmyelinated nerve fibers were vacuolated, with microfilament and microtubule disorderly arranged. Content of SP in skin tissue in D group was lower as compared with that in NC group, especially at PIW 2 [(16.8 ± 3.4) pg/g vs. (28.5 ± 5.0) pg/g, t = 2.42, P < 0.01]. There was no obvious difference in content of CGRP between NC and D groups, and also in content of glucose in skin between D and AI groups. Compared with those in D group, content of AGE in AI group at PIW 8 was decreased markedly [(27.2 ± 1.4) U/mg of hydroxyproline, t = 3.38, P < 0.05]; contents of glucose and AGE in II group at PIW 8 were significantly decreased [(1.42 ± 0.38) mg/g, (23.6 ± 1.3) U/mg of hydroxyproline, with t value respectively 1.74, 8.17, P < 0.05 or P < 0.01]. Compared with that in D group, contents of SP in AI and II groups were increased, with a delay in time of trough value. Content of CGRP showed no obvious difference among D, AI, and II groups.

CONCLUSIONSHigh glucose and accumulation of AGE are key mediators of cutaneous neuropathy and impaired wound healing in diabetes mellitus, which confirms that diabetic wound takes an atypical footing during wound repairing. Aminoguanidine and insulin can reduce contents of glucose and AGE in diabetic skin tissue, and ameliorate diabetic cutaneous neuropathy.

Animals ; Diabetes Mellitus, Experimental ; complications ; metabolism ; Glucose ; metabolism ; Glycation End Products, Advanced ; metabolism ; Male ; Peripheral Nervous System Diseases ; etiology ; Rats ; Rats, Sprague-Dawley ; Skin ; metabolism ; pathology ; Skin Diseases ; etiology ; Wound Healing

Aim To investigate the expression of Foxos in human umbilical vein endothelial cells(HUVECs)with insulin resistance(IR)induced by high glucose and high fat(HG/HF)stress and its significance.Methods First, the IR model of endothelial cells was established by HG /HF stress.The differential expression of Foxos gene in normal(Ctrl )group and HG /HF group was observed, and the subtypes with the most significant changes in Foxos were screened out, such as Foxo6.Next, Foxo6 was silenced to observe its role in endothelial cell with IR.Finally, whether the mechanism of Foxo6-mediated IR was related to the interaction of NF-κB signaling was investigated.Results The expression increase of Foxo6 was the most significant among Foxos under the IR condition induced by HG/HF.Using a small RNA interference and plasmid transfection technique, we found that the silence effect of the siRNA3 fragments targeting Foxo6 was the most significant among the siRNAs.Moreover, the further study showed that silencing the Foxo6 gene could significantly reverse the endothelial IR induced by HG/HF, and the mechanism of the reversal effect was related to the interaction between the Foxo6 and NF-κB signal.Conclusions Foxo6 mediates the endothelial cell IR induced by the HG /HF stress.The underlying mechanism is that Foxo6 can interact with NF-κBp65 and activate NF-κB signaling pathway.Silencing Foxo6 can improve the IR of vascular endothelial cells induced by HG /HF stress.

Objective: To evaluate the safety and immunogenicity of hepatitis E vaccine(HEV)in Maintenance hemodialysis(MHD)patients. Methods: Based on an open-labeled controlled trial, from May 2016 to March 2018, 35 eligible MHD patients were recruited in the Hemodialysis Center of Zhongshan Hospital Affiliated to Xiamen University as the experimental group, and 70 MHD patients with matched age, gender and underlying diseases as the control group. The experimental group received HEV at 0, 1 and 6 months according to the standard vaccination procedures, while the control group received routine diagnosis and treatment without vaccine and placebo injection to observe the safety and immunogenicity of the vaccine. The safety of vaccine in MHD population was evaluated by the incidence of adverse reactions/events in the experimental and control groups. The immunogenicity of HEV in MHD patients was evaluated by comparing the data from the phase Ⅲ clinical trial. Results: The overall incidence of adverse reactions/events was 17.1% (18/105), and there were no grade 3-4 adverse reactions/events related to vaccination. In the experimental group, the incidence of local adverse reactions/events was 20.0% (7/35), and the incidence of systemic adverse reactions/events was 17.1% (6/35).There was no significant difference in the incidence of systemic adverse reactions/events between the experimental group and the control group (P>0.05). There were 23 patients receiving 3 doses with the standard schedule. The positive rate of HEV-IgG antibody was 100% and the GMC was 14.47(95%CI:13.14-15.80) WU/ml, which showed no significant difference compared with the 46 patients in Phase Ⅲ clinical trial (t=-1.04, P>0.05). Conclusion: Recombinant HEV has good safety and immunogenicity in MHD patients.
Clinical Trials, Phase III as Topic ; Female ; Hepatitis E ; Humans ; Immunogenicity, Vaccine ; Immunoglobulin G ; Male ; Renal Dialysis ; Viral Hepatitis Vaccines/adverse effects*

Objective To explore the clinical phenotype and genetic variation of familial short stature. Method The clinical data of a familial short stature pedigree in Guangxi Zhuang autonomous region were retrospectively analyzed. The disease-causing gene was identified using targeted high-throughput sequencing combined with Sanger sequencing in May 2017. The related literature were searched and the relationship between the clinical phenotype and genotype of the ACAN gene mutation were summarized. Results The two patients were brothers, one was 9-year and 10-month old boy and the other was 7-year-old boy. Both of them had short stature. Their parents were non-consanguineous marriage and both were 150 cm in height, with. Their uncle and grandpa are also short stature. Gene sequencing revealed a novel heterozygous variation c.6193delC (p.Gln2065Serfs*27) in exon 12 of ACAN gene in both brothers, which were inherited from their father. No report of this mutation was found by searching literature and databases. A total of 11 related articles in English were retrieved. Totally (including our study) 32 patients in 41 families were reported to have the pathogenic variants of ACAN gene, including 4 variants from Chinese children, but no such reports were found in Chinese literatures. The most common clinical manifestation is idiopathic short stature, which is mostly familial but could also be sporadic. Some children also suffered from osteoarthritis, disc herniation or degeneration. Most of the children had advanced bone age, but some of them were normal or even lagged. Treatment of postponing puberty by growth hormone combined with gonadotropin-releasing hormone analogues can effectively improve final height. Conclusion Heterozygous mutation of ACAN gene can cause short stature in children and has significant familial genetic characteristics, and the clinical characteristics have no relationship with genotypes.