Development of a new cerebral ischemia reperfusion model of Mongolian gerbils and standardized evaluation system

Background:The Mongolian gerbil is an excellent laboratory animal for preparing the cerebral ischemia model due to its inherent deficiency in the circle of Willis.However,the low incidence and unpredictability of symptoms are caused by numerous complex variant types of the circle.Additionally,the lack of an evaluation system for the cer-ebral ischemia/reperfusion(I/R)model of gerbils has shackled the application of this model.Methods:We created a symptom-oriented principle and detailed neurobehavioral scoring criteria.At different time points of reperfusion,we analyzed the alteration in locomotion by rotarod test and grip force score,infarct volume by triphenyltetrazo-lium chloride(TTC)staining,neuron loss using Nissl staining,and histological charac-teristics using hematoxylin-eosin(H&E)straining.Results:With a successful model rate of 56%,32 of the 57 gerbils operated by our method harbored typical features of cerebral I/R injury,and the mortality rate in the male gerbils was significantly higher than that in the female gerbils.The suc-cessfully prepared I/R gerbils demonstrated a significant reduction in motility and grip strength at 1 day after reperfusion;formed obvious infarction;exhibited typi-cal pathological features,such as tissue edema,neuronal atrophy and death,and vacuolated structures;and were partially recovered with the extension of reperfu-sion time.Conclusion:This study developed a new method for the unilateral common carotid artery ligation I/R model of gerbil and established a standardized evaluation system for this model,which could provide a new cerebral I/R model of gerbils with more practical applications.

Tyrosinase (Tyr) Gene Mutation in Albino Mongolian Gerbil (Meriones unguiculatus)

Tyrosinase is encoded by the Tyr (c or albino) locus and is the key enzyme in pigment biosynthesis. Loss of function of this enzyme caused by gene mutation results in albinism. Most cases of albinism are caused by missense mutations of tyrosinase. Albino mutations in Tyr have been identified in various animals, including human, mouse, rat, rabbit, cattle, cat, and ferret, but not in gerbil. We created two new gerbil strains: MON/Num/a (inbred agouti phenotype) and MON/Num/c (albino phenotype). Here, we report that four nucleotide substitutions in the Tyr gene caused two missense mutations in amino acids in the albino gerbil: a G-to-A mutation at position 204 in exon 1 caused R77H, and A-to-G at position 1392 and G-to-T at position 1393 in exon 5 caused Q473R. The substitution at position 1408 in exon 5 was silent. These missense mutations are conserved in all albino phenotypes we tested. Therefore, we suggest that these mutations are responsible for albinism in gerbil.

INDUCTION OF GASTRIC INTRAEPITHELIAL NEOPLASIA OF GLANDULAR STOMACH OF MONGOLIAN GERBILS BY ELICOBACTER PYLORI

Objective： To setup an animal model of gastric carcinogenesis by Helicobacter pylori （Hp） for basic, prevention and therapeutic research of Hp-related diseases. Methods： 22 young male Mongolian gerbils were administrated with suspension of Hp strain TN2 by intragastric garage for 5 consecutive times （4×10^8 CFU/time, 1 time/4 days）. 10 male gerbils were used as negative control. Two infected gerbils were killed at 10, 20, and 30 weeks, respectively, after inoculation to monitor the development of gastric lesions. Other animals were killed at 40 experimental weeks. Pathological changes of glandular stomach were examined histologically. Results： Gastric intraepithelial neoplasias （GIN） and low-grade dysplasias were observed only in the pyloric antrum of Hp-treated gerbils （3 and 2 ones, respectively）, but not in control group （5/13 vs. 0/10, P〈0.04）. High incidence of chronic active gastritis and chronic atrophic gastritis were observed in Hp-treated animals （10/13, 76.9%）. Low incidence of chronic atrophic gastritis was also detected in negative control gerbils （3/10, 30%; P〈0.04）. Conclusion： Hp inoculation could induce chronic inflammation and malignant lesions of the glandular stomach of Mongolian gerbils conveniently.

H pylori infection causes chronic pancreatitis in Mongolian gerbils

AIM: To investigate whether chronic H pylori infection has the potential to induce pancreatitis in the Mongolian gerbil model, and whether it is dependent on an intact type Ⅳ secretion system. METHODS: Mongolian gerbils were infected with wild type (WT) H pylori typeⅠstrain B128 or its isogenic mutant B128 ?cagY (defective type Ⅳ secretion). After seven months of infection, H pylori was reisolated from antrum and corpus and H pylori DNA was analyzed by semi- nested polymerase chain reaction (PCR). Inflammation and histological changes were documented in the gastric antrum, corpus, and pancreas by immunohistochemistry. Cytokine mRNA, gastric pH, plasma gastrin, amylase, lipase, and glucose levels were determined. RESULTS: The H pylori infection rate was 95%. Eight infected animals, but none of the uninfected group, developed transmural inflammation and chronic pancreatitis. Extensive interstitial fibrosis and inflammation of the pancreatic lobe adjacent to the antrum was confirmed by trichrome stain, and immuno- histochemically. Pro-inflammatory cytokine mRNA was significantly increased in the antral mucosa of all infected gerbils. In the corpus, only cytokine levels of WT-infected animals and those developing transmural inflammation and pancreatitis were significantly increased. Levels of lipase, but not glucose or amylase levels, were significantly reduced in the pancreatitis group. H pylori DNA was detected in infected antral and corpus tissue,but not in the pancreas. CONCLUSION: H pylori infection is able to induce chronic pancreatitis in Mongolian gerbils independently of the type Ⅳ secretion system, probably by an indirect mechanism associated with a penetrating ulcer.

Short-term Celecoxib intervention is a safe and effective chemopreventive for gastric carcinogenesis based on a Mongolian gerbil model

AIM:To evaluate the optimal intervention point of a selective cyclooxygenase-2(COX-2) inhibitor,Celecoxib,for inhibiting Helicobacter pylori(H pylori)-associated gastric carcinogenesis in Mongolian gerbils(MGs).METHODS:One hundred and twelve MGs were divided into six groups(A-F).One hundred gerbils were inoculated with H pylori(groups A-E).Twelve gerbils were inoculated with vehicle broth only(group F).After 4 wk,they were given N'-methyl-N' -nitro-N-nitroso-guanidine(MNNG)(50 μg/mL) in the drinking water for 20 wk.In groups B-E,the animals were given the stock Celecoxib(10 mg/kg per day) diet from the 21st,31st,21st and 41st week respectively.The periods of administering Celecoxib were 30,20,20,and 15 wk respectively.On the 51st week,the animals were sacrificed for histological examination.Local PCNA expression was examined by the immunohistochemistry method.The expression of COX-2 protein was assessed by Western Blot.Analysis used the χ2 test.The difference was regarded as signifi cant when P value was less than 0.05.RESULTS:Seventeen percent(17/100) of H pylori-infected MGs developed gastric cancer.All of these lesions were well-differentiated adenocarcinoma.The incidence rates of adenocarcinoma in groups A-F were 40%,0%,0%,20%,25%,and 0% respectively.The inflammatory scores were higher in group B than in other groups.There was no inflammatory response noted in group F.Celecoxib treatment resulted in a significant reduction in the proliferation of H pylori-infected mucosal cells(groups B,C and D)(P < 0.01).The expression of COX-2 protein was significantly attenuated in the groups which were Celecoxib-treated for more than 20 wk(groups B,C,D).The groups treated with Celecoxib had a significantly lower rate of advanced gastric cancer(34% vs 75%,P<0.001) There were no sudden deaths in any of the groups.CONCLUSION:Short-term treatment with Celecoxib has an anti-carcinogenic effect,and resulted in less severe inflammation and inhibited the invasive degree of gastric cancer.

Anti-Helicobacter pylori therapy significantly reduces Helicobacter py/ori-Induced gastric mucosal damage in Mongolian gerbils

AIM: To investigate the effectiveness of 4 d' anti-Helicobacter pyloritherapy on the H pylori-infected Mongolian gerbils based on physiological and pathological changes.METHODS: We used 6-wk-old male gerbils orally inoculated with H pylori (ATCC43504, 2x108 CFU/mL).Seven weeks after H pylori inoculation, the animals of study group received 4 d' anti-H pylori triple therapy (H pylorieradicated group). Seven days later, all animals of the H pylori-eradicated and control groups (H pylori-infected& H pylori-uninfected groups) were sacrificed. We examined gastric mucosal lesions macroscopically, studied gastritis microscopically and determined the stomach weight ratio, myeloperoxidase (MPO) activity and prostaglandin (PG) E2 level.RESULTS: The results showed that both macroscopic and histological gastric damages were significantly less in H pylori-eradicated group than H pylori-infected group.Stomach weight ratio, MPO activity and PGE2 levels were significantly higher in H pylori-infected group than those in the other two groups.CONCLUSION: Four days' anti-H pylori therapy was effective in the improvement of H pylori-induced gastric lesions in Mongolian gerbils.

Experimental infection of Z:ZCLA Mongolian gerbils with human hepatitis E virus

AIM:To investigate whether Z:ZCLA Mongolian gerbils are readily susceptible to infection by human hepatitis E virus(HEV).METHODS:Z:ZCLA Mongolian gerbils were infected with a clinical HEV strain isolated from an acutehepati tis E patient,and virus pathogenesis was assessed in this host.Non-infected gerbils served as the control group.Feces samples from gerbils were collected weekly for reverse transcription-nested polymerase chain reaction.Serum anti-HEV Ig G and alanine aminotransferase(ALT) were detected by enzyme linked immunosorbent assay.At sacrifice,each animal's liver,spleen and kidney were collected for histopathologic examination.RESULTS:HEV-infected gerbils showed fatigue,with histopathological changes observed in the liver,spleen and kidney.HEV RNA was detected in fecal samples taken at day 7 after inoculation and the detectable levels lasted out to day 42 after inoculation.Interestingly,ALT levels were only moderately increased in the HEV-infected animals compared with the noninfected control group.CONCLUSION:Z:ZCLA Mongolian gerbils are susceptible to human HEV.

The effect of mast cell on the induction of Helicobacter pylori infection in Mongolian gerbils

INTRODUCTION Since 1982,Helicobacter pylori(Hp) has been successfully isolated and cultured,and the fact many diseases such as gastritis,peptic ulcer,gastric carcinoma

Helicobacter pyloristrain-specific modulation of gastric inflammation in Mongolian gerbils

AIM: The cag pathogenicity island (PAI) is one of potential virulence determinants of Helicobacter pylori. The Mongolian gerbil is a suitable experimental animal for the screening of virulence factors of H pylori.METHODS: Five-week-old Mongolian gerbils were inoculated with a standard H pylori strain (ATCC 43504)possessing the cag PAI or a clinical isolate lacking the genes' cluster (OHPC-0002). The animals were killed at 2, 4, 8, 24 and 48 wk after inoculation (n = 5 each), and macroscopic and histopathological findings in the stomachs were compared.RESULTS: In gerbils infected with ATCC 43504, a more severe degree of infiltration of polynuclear and mononuclear cells and lymphoid follicles was observed from 4 wk after inoculation compared to gerbils infected with OHPC-0002 especially in the antrum and transitional zone from the fundic to pyloric gland area. In addition,glandular atrophy, intestinal metaplasia, gastric ulcer and hyperplastic polyps were noted in gerbils infected with ATCC 43504, whereas only mild gastric erosions occurred in those infected with OHPC-0002.CONCLUSION: Our results indicate that the cag PAI could be directly involved in gastric immune and inflammatory responses in the Mongolian gerbils, leading to a more advanced gastric disease.

Effects of adaptation to handling on the circadian rhythmicity of blood solutes in Mongolian gerbils

The Mongolian gerbil has been widely used in many research fields and has been reported to be a diurnal laboratory animal. The circadian rhythmicity of these gerbils was investigated in the present study by measuring two hormones that show daily oscillations, cortisol and ACTH, in serum using ELISA kits. The levels of the two hormones were highest at 8:00 am and their rhythmic changes were similar to those in humans. In addition, the influence of stress of handling and blood collection on the physiological parameters of the gerbils was examined. After adaptation to handling for 1 week, some serum parameters in the animals changed. Handling and blood collection did not impact significantly on the following parameters: creatine kinase (CK), lactate dehydrogenase (LD), alanine aminotransferase (ALT), aspartate transaminase (AST), blood urea nitrogen (BUN), and albumin (ALB). However, blood glucose (GLU), total protein (TP) and globulin (GLB) significantly increased while creatinine (CRE) and albumin/globulin (A/G) significantly decreased after adaptation. This work further confirms that the Mongolian gerbil is a diurnal animal and also indicates that a suitable adaptation procedure is necessary for getting reliable results when performing experiments using these animals.

Effects of levonorgestrel-quinestrol(EP-1)treatment on Mongolian gerbil wild populations:a case study

Rodent pest population outbreaks occur frequently in grassland ecosystems in northern China.The Mongolian gerbil(Meriones unguiculatus)is a dominant pest rodent which is distributed across the semi-desert grasslands of Inner Mongolia,China.In 2009,we studied the contraceptive effect of levonorgestrel-quinestrol(EP-1),concentration 50 ppm,on a wild Mongolian gerbil population.The one-off contraceptive treatment was compared with a control group using a semi-monthly live trapping method in the Ordos Semi-desert Grassland Region of Inner Mongolia.The results show that juveniles were not recruited in spring in the treatment group.Ratios of juveniles in the control and treatment groups showed significant semi-monthly differences from spring to summer(one-way ANOVA,F_(2,14)=7.53,P<0.05).Between both groups,annual fluctuations of juvenile and total population densities were significantly different respectively(F_(2,14)=4.64,P<0.05;F_(2,18)=7.72,P<0.05).The contraceptive EP-1 delayed the normal reproductive pattern of Mongolian gerbil populations.This suppressed birth rates of gerbil populations,reduced their densities,and changed their age structures.The period of EP-1 baiting should be extended but it could be an ideal method for controlling Mongolian gerbil populations during each breeding season.

Temporal and spatial distribution characteristics in the natural plague foci of Chinese Mongolian gerbils based on spatial autocorrelation

Background:The nest flea index of Meriones unguiculatus is a critical indicator for the prevention and control of plague,which can be used not only to detect the spatial and temporal distributions of Meriones unguiculatus,but also to reveal its cluster rule.This research detected the temporal and spatial distribution characteristics of the plague natural foci of Mongolian gerbils by body flea index from 2005 to 2014,in order to predict plague outbreaks.Methods:Global spatial autocorrelation was used to describe the entire spatial distribution pattern of the body flea index in the natural plague foci of typical Chinese Mongolian gerbils.Cluster and outlier analysis and hot spot analysis were also used to detect the intensity of clusters based on geographic information system methods.The quantity of M.unguiculatus nest fleas in the sentinel surveillance sites from 2005 to 2014 and host density data of the study area from 2005 to 2010 used in this study were provided by Chinese Center for Disease Control and Prevention.Results:The epidemic focus regions of the Mongolian gerbils remain the same as the hot spot regions relating to the body flea index.High clustering areas possess a similar pattern as the distribution pattern of the body flea index indicating that the transmission risk of plague is relatively high.In terms of time series,the area of the epidemic focus gradually increased from 2005 to 2007,declined rapidly in 2008 and 2009,and then decreased slowly and began trending towards stability from 2009 to 2014.For the spatial change,the epidemic focus regions began moving northward from the southwest epidemic focus of the Mongolian gerbils from 2005 to 2007,and then moved from north to south in 2007 and 2008.Conclusions:The body flea index of Chinese gerbil foci reveals significant spatial and temporal aggregation characteristics through the employing of spatial autocorrelation.The diversity of temporary and spatial distribution is mainly affected by seasonal variation,the human activity and natural factors.

Impairment of cellular and humoral immunity in overweight Mongolian gerbils (Meriones unguiculatus)

Animal immunity is usually impaired in obesity.We know little about the effect of being overweight or obese on the immune function of wild rodents.The present study is aimed to test the hypothesis that immunity is suppressed in overweight Mongolian gerbils(Meriones unguiculatus).In the study,16 overweight(body mass:90.8-127.6 g)and 16 lean gerbils(body mass:60.5-77.7 g)were randomly selected from a total of 174 male gerbils(body mass range:55.8-144.7 g).Half of the overweight and lean males were injected with sterile saline;the others were immunochallenged(IC)with phytohaemagglutinin and keyhole limpet hemocyanin to assess cellular and humoral immunity,respectively.Body fat mass,wet and dry spleen mass,leukocyte counts,blood glucose levels and serum leptin levels were significantly higher in the overweight gerbils than in the lean gerbils.However,phytohemagglutinin response indicative of cellular immunity and immunoglobulin G concentrations was significantly lower in the IC overweight gerbils than in the IC lean gerbils.These results indicate that cellular and humoral immunity are impaired in the overweight gerbils.Excessive body fat mass,higher leukocyte counts and serum leptin levels imply that overweight gerbils are in a low grade inflammatory state.

Role of transient receptor potential vanilloid-1 in behavioral thermoregulation of the Mongolian gerbil Meriones unguiculatus

Ambient temperature considerably affects the physiology and behavior of mammals.Thermosensory and thermoregulatory abilities play an important role in the response to changing ambient temperature in endotherms.However,the molecular mechanisms of behavioral thermoregulation remain poorly understood.Transient receptor potential vanilloid-1(TRPV1)is activated by changes in ambient temperature and is involved in acute thermoregulation.Here,we aimed to determine whether TRPV1 is involved in behavioral thermoregulation in wild rodents by conducting 2 experiments.In the first,42 adult Mongolian gerbils(Meriones unguiculatus;14 per treatment)were randomly assigned to 3 housing temperatures(4,23,and 36℃for 4 weeks.In the second,20 gerbils(10 per treatment)were randomly injected with capsaicin(TRPV1 agonist)or AMG517(TRPV1 antagonist).The results showed a significant decrease in food intake and non-shivering thermogenesis in the gerbils housed at 36℃.Additionally,there was a significant increase in the preference of gerbils housed at 4℃ to low temperatures.The expression of TRPV1 protein in the brown adipose tissue(BAT)and liver was significantly positively correlated with that of protein kinase A(PKA).The expression of TRPV1 and PKA proteins in the BAT was positively correlated with the temperature preference of the gerbils.The gerbils injected with capsaicin preferred significantly lower temperatures than the control group gerbils.These findings suggest that TRPV1 and PKA are involved in behavioral thermoregulation in Mongolian gerbils.

Morphological properties of medial amygdala-projecting retinal ganglion cells in the Mongolian gerbil

The amygdala is a limbic structure that is involved in many brain functions, including emotion, learning and memory. It has been reported that melanopsin-expressing retinal ganglion cells(ip RGCs) innervate the medial amygdala(Me A). However, whether conventional RGCs(c RGCs) project to the Me A remains unknown. The goal of this study was to determine if c RGCs project to the Me A and to determine the morphological properties of Me A-projecting RGCs(Me A-RGCs). Retrogradely labeled RGCs in whole-mount retinas were intracellularly injected to reveal their dendritic morphologies. Immunohistochemical staining was performed to selectively label ip RGCs(Me A-ip RGCs) and c RGCs(Me A-c RGCs). The results showed that 95.7% of the retrogradely labeled cells were c RGCs and that the rest were ip RGCs. Specifically, Me A-c RGCs consist of two morphological types. The majority of them exhibit small but dense dendritic fields and diffuse ramification patterns as previously reported in RG_(B2)(95%), while the rest exhibit small but sparse dendritic branching patterns resembling those of RG_(B3) cells(5%). Me Aip RGCs consist of M1 and M2 subtypes. The Me A-RGCs showed an even retinal distribution patterns. The soma and dendritic field sizes of the Me A-RGCs did not vary with eccentricity. In conclusion, the present results suggest that Me A-RGCs are structurally heterogeneous. These direct RGCs that input to the Me A could be important for regulating amygdala functions.

长爪沙鼠不同感音神经性耳聋模型的建立及比较

目的利用不同的耳毒性药物建立长爪沙鼠感音神经性耳聋模型,并对3种模型进行比较分析,为感音神经性耳聋的研究提供精准的动物模型。方法选择长爪沙鼠,建立硫酸卡那霉素和呋塞米联用(KM+Fur)、新霉素(Neo)及哇巴因(Oua)3种耳毒性药物的感音神经性耳聋模型,通过听性脑干反应(ABR)和耳蜗组织形态学检测,比较分析不同耳毒性药物的损伤特点。结果KM+Fur造模后,与对照组(14.00±1.80)相比,该组(71.00±5.26)长爪沙鼠的听力阈值显著上升,耳蜗HCs大量损伤,而SGNs无明显损伤;Neo造模后,与对照组相比,40 mmol·L^(-1) Neo仅影响沙鼠高频32 kHz时听力阈值(60.00±8.66),无明显的组织学改变;而100 mmol·L^(-1) Neo组的沙鼠各频率听力阈值均显著上升,且耳蜗HCs及SGNs均严重受损;Oua造模后,1 mmol·L^(-1) Oua和10 mmol·L^(-1) Oua均可引起沙鼠各频率听力阈值显著上升,但损伤范围不同,10 mmol·L^(-1) Oua使HCs和SGNs均严重受损,而1 mmol·L^(-1) Oua仅造成耳蜗SGNs损伤。结论利用KM+Fur可建立长爪沙鼠HCs特异性受损的感音神经性耳聋模型;40 mmol·L^(-1) Neo可建立高频区听功能损伤的感音神经性耳聋模型;利用100 mmol·L^(-1) Neo和10 mmol·L^(-1) Oua可以建立长爪沙鼠HCs和SGNs均受损的感音神经性耳聋模型;利用1 mmol·L^(-1) Oua可以建立长爪沙鼠耳蜗SGNs特异性受损的耳聋模型。

唾液乳杆菌Li01对苯并芘诱导长爪沙鼠胃炎的改善作用研究

目的 建立苯并芘[benzo(a)pyrene, BaP]长时间诱导的长爪沙鼠胃炎模型,并探究唾液乳杆菌Li01对该模型的影响。方法 SPF级健康长爪沙鼠45只起初分为2组:正常对照组(NC,18只)和模型组(BaP, 27只),通过BaP灌胃造模32周后,从模型组中随机分出9只作为第3组(BaP-Li01组),用唾液乳杆菌Li01干预4周,期间记录不同组别长爪沙鼠毛发,体重等生长指标,第40周利用外周血彗星实验、流式细胞仪检测和组织病理切片探究BaP和唾液乳杆菌Li01对长爪沙鼠机体的影响。结果 经BaP灌胃后, BaP组长爪沙鼠毛发凌乱无光泽,出现耸毛、脱毛现象,而正常对照组长爪沙鼠毛发浓密光滑;另相比较于模型组,经唾液乳杆菌Li01干预后的BaP-Li01组长爪沙鼠毛发状况有所好转,接近正常对照组。与正常对照组相比,BaP组长爪沙鼠体重增长幅度明显变小, 16周后甚至小幅下降,经唾液乳杆菌Li01干预后, BaP-Li01组长爪沙鼠体重逐渐恢复。彗星实验和流式细胞凋亡检测发现, BaP摄入会造成长爪沙鼠细胞DNA损伤,而唾液乳杆菌Li01干预后可以改善、修复其损伤。病理学组织切片检查结果表明,BaP可诱发长爪沙鼠慢性胃炎,而唾液乳杆菌Li01的干预有助于胃炎的改善和修复。结论 BaP灌胃可成功建立长爪沙鼠的胃炎模型,而唾液乳杆菌Li01对其损伤有一定改善和修复作用。本研究为应用唾液乳杆菌Li01改善和修复胃炎提供了新思路。

Time-course pattern of neuronal loss and gliosis in gerbil hippocampi following mild, severe, or lethal transient global cerebral ischemia

Transient ischemia in the whole brain leads to neuronal loss/death in vulnerable brain regions. The striatum, neocortex and hippocampus selectively loose specific neurons after transient ischemia. Just 5 minutes of transient ischemia can cause pyramidal neuronal death in the hippocampal cornu ammonis (CA) 1 field at 4 days after transient ischemia. In this study, we investigated the effects of 5-minute (mild), 15-minute (severe), and 20-minute (lethal) transient ischemia by bilateral common carotid artery occlusion (BCCAO) on behavioral change and neuronal death and gliosis (astrocytosis and microgliosis) in gerbil hippocampal subregions (CA1-3 region and dentate gyrus). We performed spontaneous motor activity test to evaluate gerbil locomotor activity, cresyl violet staining to detect cellular distribution, neuronal nuclei immunohistochemistry to detect neuronal distribution, and Fluoro-Jade B histofluorescence to evaluate neuronal death. We also conducted immunohistochemical staining for glial fibrillary acidic protein and ionized calcium-binding adapter molecule 1 (Ibal) to evaluate astrocytosis and microgliosis, respectively. Animals subjected to 20-minute BCCAO died in at least 2 days. BCCAO for 15 minutes led to pyramidal cell death in hippocampal CA1-3 region 2 days later and granule cell death in hippocampal de匚tate gyrus 5 days later. Similar results were not found in animals subjected to 5-minute BCCAO. Gliosis was much more rapidly and severely progressed in animals subjected to 15-minute BCCAO than in those subjected to 5- minute BCCAO. Our results indicate that neuronal loss in the hippocampal formation following transient ischemia is significantly different according to regions and severity of transient ischemia. The experimental protocol was approved by Institutional Animal Care and Use Committee (AICUC) of Kangwon National University (approval No. KW-180124-1) on May 22, 2018.

高氧对幽门螺杆菌的清除作用及基于脂质组学的机制

目的研究高氧对幽门螺杆菌(Hp)的清除作用,并基于脂质组学探讨其可能机制。方法①高氧对Hp的清除作用。将过氧化氢300 g·L^(-1)加入生理盐水配制成含氧量为13~416 mg·L^(-1)高氧水溶液。将Hp分为Hp对照组(等体积生理盐水)、枸橼酸铋钾组(7.3 g·L^(-1))及高氧组(高氧溶液13,26,52,104,208和416 mg·L^(-1))。各组Hp置微需氧环境培养5 d后,紫外分光光度法检测菌落洗脱液在600 nm吸光度(A600 nm)值。蒙古沙鼠ig给予Hp 1×1011 CFU·L^(-1)悬液,每天1次,连续20 d,建立蒙古沙鼠Hp感染模型;选取60只感染模型沙鼠,分为Hp模型组(等体积生理盐水)、Hp模型+高氧(高氧溶液26 mg·L^(-1))组、Hp模型+三联药物组(克拉霉素8.3 g·L^(-1)、枸橼酸铋钾7.3 g·L^(-1)和替硝唑16.6 g·L^(-1)),选取20只健康沙鼠作为正常对照组(等体积生理盐水),各组分别于给药14 d后及停药14 d后处死半数沙鼠,吉姆萨染色计数胃黏膜Hp数目。②高氧对Hp细胞膜的损伤作用。将Hp分为Hp对照组(等体积生理盐水)、高氧(7,13和26 mg·L^(-1)高氧溶液)组、阿莫西林40 mg·L^(-1)组,SYTO 9/PI染色后使用流式细胞术检测Hp的存活率;BCA与ELISA法分别检测Hp对照组、高氧组胞质蛋白与DNA泄漏量;透射电镜观察高氧26 mg·L^(-1)组、阿莫西林40 mg·L^(-1)组和Hp对照组菌膜完整性。③高氧处理前后Hp脂质种类和含量变化。将Hp分为Hp对照组、高氧40 min和高氧12 h组,高效液相色谱-串联质谱法检测高氧26 mg·L^(-1)处理40 min和12 h后脂质种类及丰度,数据挖掘算法分析筛选差异脂质代谢物。结果①高氧浓度依赖性降低菌落洗脱液A600 nm值(P<0.05,P<0.01,r=-0.99)。蒙古沙鼠胃黏膜存在Hp定植,表明感染模型建立成功;Hp模型+高氧组沙鼠胃黏膜Hp数目显著低于Hp模型组(P<0.01);②高氧组Hp的存活率显著低于阿莫西林组(P<0.01);透射电镜结果显示,Hp对照组细胞膜完整、透明,胞质均匀分布。高氧组菌膜损伤、破裂、模糊不清,胞质溢出;高氧各组菌体胞质蛋白与DNA泄漏量均显著高于正常对照组(P<0.01)。③与Hp对照组相比,高氧12 h组脂质分布具有显著差异(P<0.01)。脂质组学分析筛选出2557个差异脂质,其中正、负离子模式下分别有1752和805个;正、负离子模式的上调差异脂质分别为200和85个,下调分别为73和55个,正离子模式差异脂质主要集中于脂肪酰类、甘油磷脂类和多聚乙烯类;负离子模式主要集中于脂肪酰类、固醇脂类和甘油脂类。结论高氧可清除Hp,其除菌机制与破坏Hp细胞膜的脂质结构相关。