Bilayer hydrogel dressing with lysozyme-enhanced photothermal therapy for biofilm eradication and accelerated chronic wound repair

Biofilms are closely associated with the tough healing and dysfunctional inflammation of chronic wounds. Photothermal therapy (PTT) emerged as a suitable alternative which could destroy the structure of biofilms with local physical heat. However, the efficacy of PTT is limited because the excessive hyperthermia could damage surrounding tissues. Besides, the difficult reserve and delivery of photothermal agents makes PTT hard to eradicate biofilms as expectation. Herein, we present a GelMA-EGF/Gelatin-MPDA-LZM bilayer hydrogel dressing to perform lysozyme-enhanced PTT for biofilms eradication and a further acceleration to the repair of chronic wounds. Gelatin was used as inner layer hydrogel to reserve lysozyme (LZM) loaded mesoporous polydopamine (MPDA) (MPDA-LZM) nanoparticles, which could rapidly liquefy while temperature rising so as to achieve a bulk release of nanoparticles. MPDA-LZM nanoparticles serve as photothermal agents with antibacterial capability, could deeply penetrate and destroy biofilms. In addition, the outer layer hydrogel consisted of gelatin methacryloyl (GelMA) and epidermal growth factor (EGF) promoted wound healing and tissue regeneration. It displayed remarkable efficacy on alleviating infection and accelerating wound healing in vivo. Overall, the innovative therapeutic strategy we came up with has significant effect on biofilms eradication and shows promising application in promoting the repair of clinical chronic wounds.

Engineered Bacillus subtilis alleviates intestinal oxidative injury through Nrf2-Keap1 pathway in enterotoxigenic Escherichia coli (ETEC) K88-infected piglet / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Engineered probiotics can serve as therapeutics based on their ability of produce recombinant immune-stimulating properties. In this study, we built the recombinant Bacillus subtilis WB800 expressing antimicrobial peptide KR32 (WB800-KR32) using genetic engineering methods and investigated its protective effects of nuclear factor-E2-related factor 2 (Nrf2)‍-Kelch-like ECH-associated protein 1 (Keap1) pathway activation in intestinal oxidative disturbance induced by enterotoxigenic Escherichia coli (ETEC) K88 in weaned piglets. Twenty-eight weaned piglets were randomly distributed into four treatment groups with seven replicates fed with a basal diet. The feed of the control group (CON) was infused with normal sterilized saline; meanwhile, the ETEC, ETEC+WB800, and ETEC+WB800-KR32 groups were orally administered normal sterilized saline, 5×1010 CFU (CFU: colony forming units) WB800, and 5×1010 CFU WB800-KR32, respectively, on Days 1‍‒‍14 and all infused with ETEC K88 1×1010 CFU on Days 15‍‒‍17. The results showed that pretreatment with WB800-KR32 attenuated ETEC-induced intestinal disturbance, improved the mucosal activity of antioxidant enzyme (catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx)) and decreased the content of malondialdehyde (MDA). More importantly, WB800-KR32 downregulated genes involved in antioxidant defense (GPx and SOD1). Interestingly, WB800-KR32 upregulated the protein expression of Nrf2 and downregulated the protein expression of Keap1 in the ileum. WB800-KR32 markedly changed the richness estimators (Ace and Chao) of gut microbiota and increased the abundance of Eubacterium_rectale_ATCC_33656 in the feces. The results suggested that WB800-KR32 may alleviate ETEC-induced intestinal oxidative injury through the Nrf2-Keap1 pathway, providing a new perspective for WB800-KR32 as potential therapeutics to regulate intestinal oxidative disturbance in ETEC K88 infection.

The expressions and the clinical significance of Treg and IL-35 in patients with cholangiocarcinoma / 中华肝胆外科杂志

Objective To investigate the expressions of regulatory T cell (Treg) and interleukin-35 (IL-35) in patients with cholangiocarcinoma and to explore their clinical significance.Methods Flow-cytometry,PCR,Enzyme-linked immunosorbent assay (Elisa) and immunohistochemistry were used to detect the levels of Treg and IL-35 in peripheral blood and cholangiocarcinoma tissues in 42 patients with cholangiocarcinoma.Healthy volunteers were used as a control group.Result The percentage of Treg cells to CD4 + T cells in patients with cholangiocarcinoma was (5.6 ± 1.7) ％,while that in the normal control group was (2.9 ± 0.8) ％.There was a significant difference between the two groups (P ＜ 0.05).The plasma levels of IL-35 in patients with cholangiocarcinoma was (198.4 ± 81.4) pg/ml,while that in the normal control group was (33.7 ± 18.0) pg/ml.Again,a significant difference was observed between the two groups (P ＜ 0.05).In peripheral blood mononuclear cell,the IL-35 mRNA level was positively correlated with the plasma IL-35 level (p35,R =0.795,P ＜0.05;EBI3,R =0.812,P ＜ 0.05).Immunohistochemical studies showed that FOXP3 + tumor cells and Treg cells increased significantly in tumor tissues.Conclusion Overexpressions of Treg and IL-35 in peripheral blood and tumor tissues of patients with cholangiocarcinoma suggested that they may play important roles in the development of cholangiocarcinoma.

Effects of Salvia injection on synaptic plasticity of neonatal rats with hypoxic-ischemic brain damage / 中华实用儿科临床杂志

Objective To observe the effects of Salvia injection on the brain pathology,expression of synaptophysin and the synaptic ultrastructure at different time points in neonatal rats after hypoxic-ischemic brain damage (HIBD).Methods One hundred and fifteen healthy newborn SD rats of 7-day-old were randomly divided into normal group,sham group,Salvia high dose group,Salvia low dose group,and the HIBD model group.The HIBD model was prepared by ligation of the left carotid artery combined with hypoxic environment.The rats(in normal group,sham and HIBD model groups) were injected with sterile saline[9.0 mL/(kg · d)],while the rats in high and low dose groups were injected with Salvia injection [9.0 mL/(kg · d) and 4.5 mL/(kg · d) respectively].The intraperitoneal injec tion lasted for 7 and 14 days.The rats' brains were collected at one day,7 days and 14 days after the modeling respec tively.The specimens of brain tissue were observed through hematoxylin-eosin (HE),the expression of synaptophysin (SYP) was determined by using immunohistochemistry method,and the synaptic ultrastructure in the frontal cortex was observed through transmission electron microscope.Results Finally,106 newborn rats were included in statistics analysial.(1)General observation:after HIBD modeling,the neonatal rats were mostly in the left-lateral position,with difficulties in turning over,balance abnormalities,limb shaking and other abnormal behaviors,and the delay of eyes open with the left upper eyelid ptosis.(2) Pathology:the disordered and deep stained nerve cells,the degenerated and necrotic neurons were observed in the brain tissues of the HIBD model group.(3) Expression of synaptophysin:mean density of SYP in the HIBD model group was significantly lower than the normal group on day 1,day 7,and day 14 (P ＜0.05);the SYP expression in the Salvia intervention groups increased compared with the HIBD model group(P ＜ 0.05).(4) Synaptic ultrastructure:the incomplete structure of the frontal cortex neurons,the swelling organelle and the synaptic structure damage were observed in the HIBD rats.Compared with the HIBD model group,the neuronic and synaptic ultrastructure were improved by the intervention treatment of high and low dose Salvia injection.Conclusions The mechanism of Salvia injection in treatment of neonatal rats with HIBD may be associated with the improvement in neuronal ultrastructure and synaptic reorganization.

Correlation of peripheral blood bilirubin level with Tregs in patients with bile duct stones / 中华肝胆外科杂志

Objective To study and correlate serum bilirubin and regulatory T cell (Treg) levels in patients with bile duct stone.Methods Flow-cytometry and Enzyme-linked immunosorbent assay (Elisa) were used to study the peripheral blood expression level of Tregs and the bilirubin level in 27 patients with bile duct stones and jaundice.The changes in the expression level of Tregs and the bilirubin level were studied and correlated before and after treatment.Results After treatment,both the peripheral blood bilirubin level,the Tregs expression level and the cell cytokines decreased significantly.The total bilirubin level decreased from (102.8 ± 33.1) mmol/L to (15.3 ± 5.7) mmol/L (P ＜ 0.05),the direct bilirubin level decreased from (38.1 ± 12.8) mmol/L to (5.0 ± 1.6) mmol/L (P ＜0.05);the percentages of CD4+ CD25 +Foxp3 + T cells in CD4+ T decreased from (4.2 ± 2.0) ％ to (2.4 ± 1.0) ％ (P ＜ 0.05).Before treatment,the levels of IL-10 and TGF-β were 171.4 ± 13.7 and 2016 ±657 pg/ml but after treatment,the two cytokines decreased to 92.1 ± 7.4 and 1 686 ± 168 pg/ml,respectively (P ＜ 0.05).Conclusions Patients with bile duct stones and jaundice presented with high expressions of bilirubin and Tregs level.These expressions returned to normal after effective treatment.The Tregs expression level was positively correlated with the bilirubin level.

Shape-memory polymer based on D,L-poly(lactic acid):Biosecurity and cytocompatibility / 中国组织工程研究

BACKGROUND:Previous experiments have prepared shape memory polymer based on D,L-poly(lactic acid). According to domestic technical requirements for biological y evaluating biomaterials and medical equipments, tissue-engineered grafts must be subjected to preclinical experiment for biosecurity and cytocompatibility evaluation. OBJECTIVE:To observe the biosecurity of the shape memory polymer based on D,L-poly(lactic acid). METHODS:(1) Bacterial endotoxin test:polymer extract, endotoxin working standard solution and checking standard water were added into limulus reagent, respectively. (2) Sensitization test:Polymer extract+Freund’s complete adjuvant+physiological saline and Freund’s complete adjuvant+physiological saline were injected into the scapula of Kunming mice. After induction by intradermal injection, local induction and excitation, stimulate skin erythema and edema degree were observed in animals. (3) Acute toxicity test:Kunming mice received intraperitoneal injection of 100%, 50%, 25%polymer leaching solution and physiological saline, respectively. (4) 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay for cel proliferation:The direct method was that human umbilical cord vascular endothelial cel s were inoculated onto the polymer film, lactic acid and glass, respectively;the indirect method was that human umbilical cord vascular endothelial cel s were inoculated into the polymer leaching solution, acrylamide solution and 1640 culture solution. RESULTS AND CONCLUSION:This shape memory polymer based on D,L-poly(lactic acid) is free of bacterial contamination in compliance with the biosecurity standards, and it has no al ergenic and toxicity but has good cytocompatibility.

Characterization and Diagnostic Use of a Monoclonal Antibody for VP28 Envelope Protein of White Spot Syndrome Virus / 中国病毒学·英文版

The gene encoding the VP28 envelope protein of White spot syndrome virus (WSSV)was cloned into expression vector pET-30a and transformed into the Escherichia coli strain BL21.After induction,the recombinant VP28 (rVP28) protein was purified and then used to immunize Balb/c mice for monoclonal antibody (MAb) production.It was observed by immuno-electron microscopy the MAbs specific to rVP28 could recognize native VP28 target epitopes of WSSV and dot-blot analysis was used to detect natural WSSV infection.Competitive PCR showed that the viral level was approximately 104 copies/mg tissue in the dilution of gill homogenate of WSSV-infected crayfish at the detection limit of dot-blot assay.Our results suggest that dot-blot analysis with anti-rVP28 MAb could rapidly and sensitively detect WSSV at the early stages of WSSV infection.

Effects of vagus nerve stimulation on seizure in animals with epilepsy / 中国组织工程研究

BACKGROUND: Vagus nerve stimulation (VNS) is a neurophysiological therapy (NPT) of refractory epilepsy, which can control the seizure by stimulating the vagus nerve stem in cervical part.OBJECTIVE: To study the effect of intermittent left-side VNS on seizure of epileptic animals, and provide theoretic basis for the interaction of somatic information and that of internal organs.DESIGN: Observation study.SETTING: Department of Neurobiology, Capital University of Medical Science.MATERIALS: The experiment was performed in the Laboratory of Electrophysiology in Department of Neurobiology, Capital University of Medical Science from March 2000 to September 2002. Thirty-four healthy adult SD rats and 8 rabbits, weighting (220-250) g and (2.2-2.5) kg respectively were selected.METHODS: ①Ten rats were intramuscularly injected with (150 000-160 000) U of penicillin (PCN). VNS effects on epileptiform activities of rats were studied by observing the changes in electrocorticogram (ECoG)and behavior of rats before and after VNS.②(0.24-0.48) mg of PCN was injected into the hippocampus of another 8 rabbits to induce epilepsy, and VNS effects on ECoG of epileptic rats were observed. ③Seizures of 16 rats were induced by Kainic acid, and changes in discharge activity of hippocampal neuron, ECoG and behavior of epileptic rats were observed after VNS. ④Seizures of 8 rabbits were induced by cortical injection of strychnine with microinjector, and VNS effects on ECoG of rabbits with epilepsy induced by acute cortical injury were observed.MAIN OUTCOME MEASURES: ①VNS effects on seizure of rats with epilepsy induced by PCN. ②VNS effects on seizure of rats with epilepsy induced by Kainic acid. ③VNS effects on epileptiform ECoG of rabbits with epilepsy induced by strychnine.RESULTS: A total of 34 rats and 8 rabbits were involved in the analysis of results. VNS could remarkably suppress the seizure of epileptic animals,and epileptiform ECoG, epileptiform discharges of hippocampal neuron and behavior significantly changed with the total effective rate greater than 50%. The total effective rate of VNS before seizure was greater than 80%.In epilepsy group indoeed by intramuscular injection of PCN, ECoG and behavior were markedlly aneliorated respectively for 40% and 50% of rats.In epilepsy group induced by injection of PCN in hippocampus, the ECoG was siguificantly ameliorated in 50% rats. In epileptic rabbit group induced by partial injection of strychnine via cerebral cortex, the epilepti form wave iu ECoG was controlled by VNS in 50 % of animals.CONCLUSION: VNS can effectively suppress seizure of epileptic animals. The antiepileptic effect of VNS is associated with cerebral cortical aud hippcampal neurons. Somatic epileptiform activity could be effectively inhibited by the integration of visceral afferent information in cortical and hippocampal parts.

Effect and mechanism of vagus nerve stimulation on seizure in animal with epilepsy / 中国康复理论与实践

@#ObjectiveTo study the effect and mechanism of vagus nerve stimulation(VNS) on seizure in animal with epilepsy.MethodsSeizures of 34 rats and 8 rabbits were induced by Penicillin, Kainic acid(KA) and Strychnice respectively. Electrocorticographic(ECoG), electrical activition of hippocampal neurons and behaviour were observed to evidence the effects of left intermittent VNS .ResultsVNS could suppress seizures in animal models with epilepsy. There were significant changes in epileptiform ECoG, discharges of hippcampal neuron and behaviour. ConclusionSomatic seizure can be effectively inhibited by visceral afferent inputs through integration in cortical and hippocampal parts.

Imaging Diagnosis of Testicular and Epididymal Tumors or Inflammatory Nodes in Children / 实用放射学杂志

Objective To analyzed imaging features of testicular and epididymal tumors or inflammatory nodes in children and to improve its diagnostic accuracy.Methods 13 cases underwent ultrasonography,of them,plain CT scan in 10 cases,enhanced CT scan in6 cases.Its signs were retrospectively analyzed and compared with surgical and pathological results.Results On ultrasonography,theinhomogenous and different echo could be viewed in 13 cases,color Doppler flow imaging was abundant in 8 masses,3 cases with littler,empty blood flow in 2 cases and retroperitoneal lymphatic metastasis were viewed in 2 cases.On plain CT scan,masses were mixed density in 4 cases,calcification could be seen within tumor in 3 cases,masses were soft tissue or main soft tissue density in 6 cases.Contrast-enhancedCT scan displayed obviously and inhomogenously in 4 cases,lightly enhancment in 1 case and no enhanced in 1 case.By surgical andpathological confirmed,3 were mature treatomaes,1 was immaturity teratoma,4 were yolk sactumor,3 were inflammatory nodes,1 wasrhabdomyosarcoma and 1 was cystic lymphangioma.Conclusion Each kind of testicular tumors in children has its owns CT and UScharacteristics.In combination of CT and US can carry high diagnostic accuracy.