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Objective:To investigate the effect of KRT5 knockdown in keratinocytes on melanin content in co-cultured melanocytes, and to explain mechanisms underlying formation of hyperpigmented lesions in reticulate pigmented anomaly of the flexures (Dowling-Degos disease, DDD) .Methods:HaCaT cells with heterozygous mutations in the KRT5 gene were obtained by using clustered regularly interspaced short palindromic repeats (CRISPR) -CRISPR-associated protein 9 (Cas9) technology (experimental group) , and HaCaT cells transfected with non-targeting single guide RNA:Cas9 protein complex served as control group, both of which were in vitro co-cultured with primary human melanocyte cells (HEMn) separately. Immunofluorescence study was conducted to determine the expression of cytokeratin and melanosomes in co-cultured cells; melanin content was detected in melanocytes in different co-culture groups, which were obtained by differential trypsinization. Immunohistochemical study was performed to determine the expression of melanocyte-specific premelanosome protein 17 (Pmel17) in skin lesions in a patient with DDD carrying a KRT5 mutation and normal skin tissues in a healthy control. Results:Sanger sequencing showed a heterozygous mutation (c.1delA) at the initiation codon of exon 1 of the KRT5 gene in HaCaT cells in the experimental group, but no mutation in the KRT5 gene in the control group. Western blot analysis showed that the KRT5 protein expression was significantly lower in the experimental group (0.60 ± 0.05) than in the control group (1.00 ± 0.00, t = 32.38, P = 0.001) . Compared with the co-culture system in the control group, the number of Pmel17-labeled melanosomes markedly increased with the melanin content elevated by 52.5% ( t = -3.48, P = 0.025) in the HEMn cells co-cultured with HaCaT cells in the experimental group. Immunohistochemical study showed that the Pmel17 expression increased in the skin lesions in the DDD patient with KRT5 mutation compared with the normal skin tissues in the healthy control. Conclusion:The effect of HaCaT cells with CRISPR-Cas9-induced KRT5 mutation on the co-cultured HEMn melanocytes was verified by the successfully established in vitro co-culture system, which provides a primary cell model for further studies on interaction mechanisms between keratinocytes and melanocytes, and on pathogenesis of skin pigmentation abnormalities.
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Objective:To analyze and compare VP1 and VP4 genetic characteristics of human enterovirus 71(EV71) isolated from children with severe and mild hand-foot and mouth disease(HFMD).Methods:EV71 strains isolated from severe HFMD patients (6 cases) and mild HFMD patients (6 cases) in Taizhou district, Zhejiang province were included during 2016.Total virus RNA was extracted by Viral RNA Mini Extraction Kit, and reverse transcription polymerase chain reaction(RT-PCR) was used to amplify the sequence of the VP1 and VP4 genes of EV71.And then the sequencing results were compared with those of A, B, C genotype reference EV71 strains from GenBank by nucleotide alignment and amino acid alignment analysis.Results:There were no statistically significant differences in sex and age between the two groups(χ 2=14.51, t=2.82, all P<0.05). The homogeneity between EV71 strains from severe patients and mild patients was 95.8%~99.6% and 99.1%~100.0% for VP1 nucleotide sequences and amino acid sequences, respectively, while 95.0%~99.9% and 99.0%~100.0% for VP4 nucleotide sequences and amino acid sequences, respectively.The twelve EV71 strains isolated from HFMD patients in Taizhou shared the highest identity with EV71-genotype C, especially genotype C4a.In addition, compared with the mild patients, three respective strains from severe HFMD patients showed mutations at the residue 170 in the VP1 protein(V→L), residue 293 in the VP1 protein(A→S) and residue 7 in the VP4 protein(T→A). Conclusion:All EV71 strains isolated from severe and mild HFMD patients in Taizhou district share high homology of nucleotide and amino sequence, and all of them belong to subgenogroup C4a.The mutations in the VP1 and VP4 of EV71 might be related to HFMD disease severity.
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Psoriasis is a chronic recurrent inflammatory skin disease,and its pathogenesis is still unclear.At present,it is believed that immune,genetic and environmental factors play important roles in the occurrence of psoriasis.More and more studies have indicated that the imbalance of intestinal flora is associated with immune diseases,especially inflammatory bowel disease.The pathogenesis and clinical manifestations of psoriasis are similar to those of inflammatory bowel disease,and psoriasis is proved to be associated with changes in intestinal microbial flora.This review discusses the association between the imbalance of intestinal flora and psoriasis.
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Objective To explore the clinical efficacy,adverse reactions and serum levels of IL -10 and TNF-αin children with peptic ulcer undergoing sequential therapy.Methods 68 children with peptic ulcer were selected and randomly divided into two groups,34 cases in each group.The control group received quadruple therapy, namely omeprazole,amoxicillin,clarithromycin and bismuch treatment for seven days.The treatment group underwent sequential therapy:the first 5 d of oral omeprazole,amoxicillin treatment,and the next 5 d omeprazole,amoxicillin and inidazole treatment.The clinical efficacy,adverse reactions,IL-10 and TNF-αlevels of the two groups were com-pared.Results The total effective rates after treatment of the control group and treatment group were 88.24% and 91.18% respectively,there was no statistically significant difference between the two groups(χ2 =1.21,P>0.05). After treatment,the levels of IL-10 and TNF-αin the control group were (24.93 ±6.29)pg/mL and (37.93 ± 8.28)pg/mL,which were significantly decreased (t=5.52,P<0.05,t=8.51,P<0.01).And the levels of IL-10 and TNF-αin the treatment group were (21.36 ±6.31)pg/mL and (29.67 ±6.38)pg/mL,which were significantly decreased(t=11.15,12.29,all P<0.01).The levels of IL-10 and TNF -αof the treatment group were much significant than those of the control group after the sequential therapy (t=3.32,P<0.05,t=8.71,P<0.01). Conclusion Sequential therapy for the treatment of children with peptic ulcer has better effect than the quadruple therapy,and can reduce serum IL-10 and TNF-αlevels,it is worthy of promoting.
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Objective To investigate the change of blood lactate level in children with severe sepsis,and its relationship with clinical prognosis.Methods 90 children with severe sepsis who treated in our hospital from February 201 3 to May 201 4 were selected as the study subjects.According to the prognosis of children,they were divided into survival group and death group,45 cases in each group.The blood lactate levels at different time points, blood lactate clearance rates between the two groups at different time points as well as fibrin,oxygenation index and D -dimer levels were compared after admission.Results After treatment,the fibrin,oxygenation index and D -dimer levels in the two groups were improved.The fibrin and D -dimer levels in the survival group[(2.71 ±0.31 )ng/mL, (0.89 ±0.1 0)mg/L)]were lower than those in the death group[(2.89 ±0.21 )ng/mL,(1 .26 ±0.1 8)mg/L)],the differences were significant(t =3.224,P =0.001 ;t =1 2.053,P =0.000).The oxygenation index of the survival group[(1 96.23 ±1 4.69)mmHg)]was higher than that of the death group [(1 80.23 ±21 .03 )mmHg)],the difference was significant(t =4.1 84,P =0.000).The EGOT compliance rate,APACHE Ⅱscore and MODS incidence rate of survival group were significantly lower than those of the death group,the differences were significant(t =7.200,P =0.007;t =9.1 49,P =0.000;t =29.298,P =0.000).The blood lactate levels at each time points in the survival group were significantly lower than the death group,the differences were statistically significant(t =50.543, P =0.000;t =33.932,P =0.000;t =1 7.91 5,P =0.000;t =28.703,P =0.000).The 6 h,24 h blood lactate clearance rates≥1 0% of the survival group (73.33%,80.00%)were significantly higher than those of the death group(37.78%,44.44%),the differences were significant(χ2 =1 1 .520,P =0.000;χ2 =1 2.1 00,P =0.000). Conclusion Lactate level in children with sepsis is an important indicator of prognosis in children with severe sepsis,with guidance for the treatment of children with sepsis.
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Objective To compare the clinical effect of intraosseous versus traditional infusion in critical pediatric patients under resuscitation.Methods 56 critical pediatric patients under resuscitation,who were difficult to establish a venous access,were divided into two groups:intraosseous infusion group (IOI group)and traditional group.The clinical characteristics,lab tests,clinical efficacy and complications of all patients were recorded in detail. Results The general characteristics of two groups were comparable.The mean operation time to establish an emergency infusion access in the IOI group was (1.01 ±0.53)min,which was significantly shorter than (8.97 ±2.36)min of the traditional group(t =4.856,P <0.01).The IOI group had a greater efficacy over the traditional group (50.0% vs 36.7%),in the IOI group,the patients showed obvious positive effect and less patients had no efficacy(χ2 =18.476, P <0.05 ).In addition,lower complication rate and mortality were observed in the IOI group.Conclusion For critical pediatric patients under resuscitation whose intravascular access cannot be achieved through peripheral venous,intraosseous infusion is prior to traditional infusion.
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Objective To investigate the clinical effect of L -glutamine gung shimron acid sodium in combi-nation with esomeprazole in the treatment of pediatric peptic ulcer and its effects on immunological function and intes-tinal flora.Methods 120 patients of pediatric peptic ulcer meeting the inclusion criteria were randomly divided into two groups equally.Besides conventional treatments,the control group was treated with esomeprazole,and the observa-tion group was treated with L -glutamine gung shimron acid sodium in combination with esomeprazole.After the course of treatment,the curative efficacy was compared after the eight -week follow -up.Before and after the treat-ment,T lymphocyte subpopulation (CD +4 ,CD +8 ,CD +4 /CD +8 )reflecting cellular immune function,intestinal flora and adverse reactions were compared.Results After the eight -week follow -up,the total effective rate of the observa-tion group was 96.7%,which was significantly higher than 86.7% of the control group (χ2 =3.93,P <0.05).After treatment,levels of CD +4 [(43.5 ±6.6)% vs (40.9 ±6.2)%],CD +4 /CD +8 [(1.72 ±0.38)vs (1.38 ±0.45)] were statistically higher and level of CD +8 [(26.2 ±3.7)% vs (30.1 ±5.0)%]was statistically lower in the obser-vation group in comparison with the control group (t =2.22,P <0.05;t =4.47,t =4.86,P <0.01).As to intestinal flora,the observation group had statistically higher concentration of lactobacillus [(7.5 ±1.3 )LonN /g feces vs (5.8 ±1.2)LonN /g feces],bifidobacterium[(8.3 ±1.1)LonN /g feces vs (6.6 ±1.4)LonN /g feces]in the feces in comparison with the control group after treatment (t =7.44,t =7.40,both P <0.05).During the treatment,there were no severe adverse reactions.Conclusion L -glutamine gung shimron acid sodium in combination with esome-prazole is effective for pediatric peptic ulcer,which can significantly increase curative efficacy,enhance cellular immune function and effectively regulate intestinal flora with minor adverse reactions.
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Objective To analyze the therapeutic effect of qiweiqingyan aerosol combined with ribavirin in the treatment of herpetic angina of children hand-foot-mouth disease,and to compare the clinical effects with sim-ple use of ribavirin.Methods 160 children with hand foot and mouth disease were divided into two groups by random number table.They were the observation group (80 cases)and control group (80 cases),respectively.Two groups were both given ribavirin aerosol in the treatment,the observation group were added with qiweiqingyan aerosol agent to carry out treatment,then the clinical curative effects of the observation group and control group were copared.Results In the observation group,the total effective rate was 96.25%,and the total effective rate of the control group was 85.00%,the difference between the two groups had statistical significance (χ2 =5.959,P<0.05).Marked effective rate of the observation group was 86.25%.In the control group,the significant efficiency was 37.50%.The difference between the two groups had statistical significance (χ2 =40.300,P<0.05 ).Defervescence time and bleb disappear time were shorter in the observation group compared to the control group,the difference between the two groups was statistically significant (t=47.880,8.063,5.100,all P<0.05).The adverse reactions in the observation group were significantly lower than those in the control group (χ2 =9.608,P<0.05 ),after statistical analysis.Conclusion Using qiweiqingyan aerosol combined with ribavirin in the treatment of children hand foot and mouth herpangina,can significantly shorten the time of treatment,reduce adverse reactions and improve the cure rate.
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Objective To explore an accurate,objective and simple method for barrel distortion correction and lesion area measurement by assistance of computer.Methods The software of Panaroma tool was employed to correct barrel distortion of endoscopy and Image J to measure lesion size and manage the relative measurement error.Computed measurement in vitro of lesion area was established,firstly,by identification of correction factor of Panorama tool to minimize measurement error; then by determination of influence of object distance change between the lens and the image.This measurement was used on patients with gastric ulcer for focal area.Results were compared with those of traditional method.Results Number of 0.1was determined to be the correction factor for barrel distortion of endoscopy.Prior to the correction of the barrel distortion,the relative error of measurement gradually increased with the increasing distance between endoscopy and the image.However,different object distances did not exert influence on the relative measurement error when barrel distortion was corrected by Panaroma tool ( P =0.141 ).A total of 50 foci of gastric ulcer were measured,results from combinational treatment of Panorama tool and Image J showed the areas (35.0 ± 5.0) mm2 were significantly larger than those determined by traditional method [ ( 29.1 ± 4.1 ) mm2,P =0.000 ],with a correlation coefficient of 0.988.Conclusion Computed endoscopic lesion measurement is a relatively accurate,objective and simple method to determine the area of gastric lesions.