Objective To explore the mechanisms underlying the mediating effect of regulatory B (Breg)cells in the pathogenesis of pemphigus.Methods Peripheral blood specimens were collected from 48 patients with pemphigus at different degrees of severity and 20 healthy controls.Flow cytometry was conducted to analyze the frequency of Breg(CD19 +CD24hiCD38hi)cells in peripheral CD19+ B cells,enzyme linked immunosorbent assay(ELISA)to determine the titer and subtypes of anti-desmoglein(Dsg)1 and 3 antibodies in sera from these subjects.The relationship of anti-Dsg 1 and 3 antibodies with the frequency of Breg cells was assessed by Spearman's rank correlation test.Results Significant differences were observed in the proportion of Breg cells in CD19+ B cells between patients with pemphigus and healthy controls(11.54％ ± 0.97％ vs.8.19％ ± 0.85％,P =0.04),as well as between patients with acute mild pemphigus and those with moderate and severe pemphigus(5.17％ ± 2.14％ vs.11.38％ ± 5.30％ and 11.17％ ± 5.31％,P =0.042,0.046,respectively).The frequency of Breg cells was positively correlated with the absorbance value for anti-Dsg1 antibodies of IgG4 subclass and IgG4/IgG1 ratio(r =0.527,0.565,respectively,both P ＜ 0.01).Conclusions Breg cells are mainly associated with the production of antibodies in and disease severity of pemphigus,while the actual mechanism of action remains unknown.

Objective To assess the correlation between the subclasses of antibodies against desmoglein (Dsg) 1 and Dsg3 and disease activity in patients with pemphigus. Methods Sera were collected from 47 patients with pemphigus, and ELISA was performed to determine the titers and subclasses of antibodies against Dsg1 and Dsg3. The correlation of antibody titers and subclasses with disease activity was assessed.Results Clinical phenotype was associated with antibody profiles in these patients with pemphigus. Of 17 patients with mucocutaneous involvement, 14 (82.4%) had both anti-Dsg3 and anti-Dsg1 antibodies; of 16 patients with cutaneous involvement, 15 (93.7%) had anti-Dsg1 antibody, only 1 (6.3%) developed anti-Dsg3 antibody; of 6 patients with mucosal involvement, all ( 100% ) had only anti-Dsg3 antibody. The serum levels of antibodies against Dsg1 and Dsg3 were increased, but not in parallel with the disease severity in these patients.Moreover, the subclasses of antibodies were correlated with disease severity. IgG4 subclass antibodies against Dsg1 and Dsg3 predominated in patients with pemphigus at active stage, whereas IgG1 subclass in those at remission stage. The serum levels (expressed as absorbance value) of IgG4 and IgG1 subclass antibodies were 1.92 ± 1.21 and 0.60 ± 0.61 respectively, with the ratio of IgG4 to IgG1 more than 1, in patients with antiDsg1 antibodies at active stage, 0.03 ± 0.02 and 0.22 ± 0.11 respectively with the ratio of IgG4 to IgG1 less than 1, in those at remission stage, 2.35 ± 2.17 and 1.84 ± 1.16 respectively with the ratio of IgG4 to IgG1 more than 1 in patients with anti-Dsg3 antibodies at active stage, 0.15 ± 0.16 and 1.05 ± 0.77 respectively with the ratio of IgG4 to IgG1 less than 1 in those at remission stage. Conclusions The subclasses of pemphigus-specific antibodies are closely correlated with the disease severity in patients with pemphigus. The detection of subclasses and titers of anti-Dsg1 and anti-Dsg3 antibodies may aid in the diagnosis of and monitoring of disease severity in pemphigus.

ObjectiveTo investigate the association between the Clock T3111C and T257G gene polymorphisms and sleep epilepsy patients in Han population of Hunan province.MethodsThree hundred and eleven subjects with epilepsy ( sleep epilepsy group ( n =112 ),aperiodic group ( n =95 ),awakening epilepsy group ( n =104 ) ) and 300 sex- and age-matched healthy controls were enrolled in the study.Genomic DNAs were extracted from peripheral blood leucocytes by phenol-chloroform methods.The Clock T3111C and T257G polymorphisms were detected by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP).Results(1) Two genotypes(TT and TC) were detected in Clock T3111C.The frequency of Clock site T3111C genotypes in all of the people was 86.09％ (TT,526/611),13.91％(TC,85/611),0( CC),T allele gene frequency was 93.04％ (1134/1222) and C allele gene frequency was 6.96％ (85/1222).There was no significant difference in genotype and gene distribution of Clock gene T3111C polymorphism between sleep epilepsy group,aperiodic group,awakening epilepsy group and control group.(2)Two genotypes(TT and TG) were detected in Clock T257G.The frequency of Clock site T257G genotypes in all of the people was 85.92％ (TT,525/611 ),14.08％ (TG,86/611 ),0( GG),T allele gene frequency was 92.96％ (1136/1222) and G allele gene frequency was 7.04％ (86/1222).There was no significant difference in genotype and gene distribution of Clock gene T257G polymorphisms between sleep epilepsy group,aperiodic epilepsy group,awakening epilepsy group and control group.(3)There was an almost complete correspondence (complete linkage disequilibrium) of bases between the positions 257 and 3111.ConclusionClock gene T3111C and T257G polymorphisms are not associated with sleep epilepsy in Han population of Hunan province.

Objective:To evaluate the role of miR-146a in hippocampal inflammatory responses in postoperative cognitive dysfunction (POCD) in mice.Methods:One hundred and sixty clean-grade male C57BL/6 mice, aged 12-16 weeks, weighing 22-28 g, were divided into 5 groups ( n=32 each) using a random number table method: control group (group C), group POCD, miR-146a agomir group (group Ag), miR-146a antagomir group (group At) and negative control group (group NC). The mice were subjected to an intramedullary fixation for tibial fracture under 1.5% isoflurane anesthesia to establish POCD model.At 2 days before operation, miR-146a agomir 0.5 nmol (0.1 nmol/μl) was injected into bilateral hippocampi in group Ag, miR-146a antagomir 2.5 nmol (0.5 nmol/μl) was injected in group At, miR-146a negative control solution 2.5 nmol (0.5 nmol/μl) was given in group NC, and the animals in group C did not receive any treatment.At 1 day before operation and at 1, 3 and 7 days after operation, open-field test was performed to evaluate spontaneous motor activity, and contextual fear conditioning test was performed to evaluate cognitive ability 15 min later.At 1 and 3 days after operation, the animals were sacrificed and hippocampi was removed for determination of expression of CD11b (a marker for activation of microglia) in hippocampal CA1 region by immunofluorescence staining.At 6, 12 and 24 h after operation, the expression of miR-146a was detected by quantitative real-time polymerase chain reaction, the expression of interleukin-1 receptor-associated kinase 1 (IRAK1), tumor necrosis factor receptor-associated factor 6 (TRAF6), nuclear factor kappa B p65 (NF-κB p65) and tumor necrosis factor-alpha (TNF-α) was determined by Western blot and interleukin-1beta (IL-1β) and IL-6 contents were determined by enzyme-linked immunosorbent assay. Results:There was no significant difference in the total exploring distance in the open-field test or percentage of freezing time in tone-fear conditioning test at each time point among the five groups( P>0.05). Compared with group C, the percentage of freezing time in the contextual fear conditioning test was significantly decreased at 1, 3 and 7 days after operation, the expression of CD11b at 1 and 3 days after surgery and expression of miR-146a, IRAK1, TRAF6, NF-κB p65 and TNF-α were up-regulated and the contents of IL-1 β and IL-6 were increased at 6, 12 and 24 h after operation in group POCD ( P<0.05). Compared with group NC, the percentage of freezing time in the contextual fear conditioning test was significantly increased at 1, 3 and 7 days after operation, and the expression of CD11b was down-regulated at 1 and 3 days after surgery, and the expression of miR-146a, IRAK1, TRAF6, NF-κB p65 and TNF-α was up-regulated and IL-1β and IL-6 contents were decreased at 6, 12 and 24 h after operation in group Ag, and the percentage of freezing time in the contextual fear conditioning test was decreased at 1, 3 and 7 days after operation, the expression of CD11b at 1 and 3 days after surgery was up-regulated, the expression of miR-146a was down-regulated and IRAK1, TRAF6, NF-κB p65 expression was up-regulated at 6, 12 and 24 h after operation, TNF-α expression was up-regulated and IL-1β and IL-6 contents were increased at 12 and 24 h after operation in group At ( P<0.05). Conclusion:miR-146a is involved in the process of hippocampal inflammatory responses, and the mechanism may be related to the inhibition of IRAK1-TRAF6-NF-κB signaling pathway in mice.

Objective To summarise the imaging features of rare mastitis and explore the diagnostic value of ultrasound, mammography and MRI for rare mastitis. Methods The record of 24 patients diagnosed as rare mastitis in our hospital from Jan. 2000 to Jun. 2009 was reviewed, including clinical manifestations, pathological results, imaging diagnosis and diagnostic accurate rate. Results Of the 24 patients, 14 patients were ductal ectasia with chronic mastitis, 3 granulomatous mastitis, 6 chronic abscess and 1 mammary tuberculosis. 13 patients underwent ultrasonic scan, 12 patients underwent mammography and 3 patients underwent MRI, with the diagnostic accurate rate 77%, 25% and 100% respectively. Conclusions There are no special imaging manifestations for most rare mastitis, however, some differential characteristics still exist. MRI has a higher accuracy compared to ultrasound and mammography. The combination of multiple imaging methods can improve diagnostic accuracy.

Objective To investigate the changes of miR-146a expression in serum,hippocam-pus and prefrontal cortex in postoperative cognitive dysfunction (POCD) mice.Methods Fifty healthy male C57BL/6 mice,aged 12-14 weeks,weighing 25-30 g,were randomly divided into two groups (n =25 each)using a random number table:control group (group C)and anesthesia plus sur-gery group (group AS).Mice in group AS underwent open tibial fracture of the left hind paw with in-tramedullary fixation in aseptic conditions under general anesthesia with 2.1% isoflurne.Ten mice in each group received the fear conditioning test (FCT)on the 1,3 and 7 days after anesthesia/surgery. The rest of mice were sacrificed 24 h before (baseline),and 6,12,24,48 h after anesthesia/surgery, and then the serum,prefrontal cortex and hippocampus were collected or removed for detection of the expression of miR-146a using quantitative reverse transcription polymerase chain reaction (qRT-PCR).Results Compared with group C,the percentage of freezing time in contextual FCT was sig-nificantly decreased (P <0.05)in group AS,while no significant change in freezing time percentage was found in tone-cued FCT.In serum,compared with group C,miR-146a expression at 6,12,24, 48 h after anesthesia/surgery was significantly up-regulated in group AS (P < 0.05 );and in group AS,the expression of miR-146a was significantly decreased 6,24,48 h as compared to that at 12 h after anesthesia/surgery (P <0.05).In hippocampus,compared with group C,miR-146a expression at 6,12,24,48 h after surgery was significantly up-regulated in group AS (P <0.05);and in group AS,the expression of miR-146a at 6,48 h after surgery was significantly decreased as compared to that at 12 h after anesthesia/surgery (P <0.05).In prefrontal cortex,compared with group C,miR-146a expression at 24,48 h after surgery was significantly up-regulated in group AS (P <0.05);and in group AS,the expression of miR-146a was significantly increased at 48 h as compared to that at 24 h after anesthesia/surgery (P <0.05).Conclusion The expression of miR-146a in serum,hippocam-pus and prefrontal cortex in POCD mice was up-regulated,and changes of miR-146a expression may be related to the development of POCD.

Objective To investigate the effectiveness of three different anesthetic techniques in intraventricular catheterization and its effect on the survival rate of rats. Methods Thirty Wistar rats were equally allocated into 3 groups:chloral hydrate group,pentobarbital sodium group and isoflurane group. Intraventricular catheterization was performed in the rats after anesthesia with i. p. injection of chloral hydrate and pentobarbital sodium, and isoflurane inhalation, respectively. Levels of blood glucose were detected before and at 15 and 30 minutes and 1, 3, 7, 14, 28 days after anesthesia. Body mass and 24-hour food intake were recorded before and at 1, 3, 7 days after anesthesia. The onset time and effective time of anesthesia, operation time and the survival rates on 30 days of the rats were compared and analyzed. Results The onset time and effective time of anesthesia, and the operation time in the isoflurane group were shorter than that in the chloral hydrate group, while these parameters in this group were shorter than that in the pentobarbital sodium group. Blood glucose in the chloral hydrate group was apparently increased during the surgical operation, while the body mass, 24-hour food intake and blood glucose were decreasing since one day after operation, and all the rats in this group died during the 30-day observation, mainly, due to enteroplegia. Blood glucose in the pentobarbital sodium group was mildly increased after anesthesia, while the body mass, 24-hour food intake and blood glucose were mildly decreased at one day after operation and recovered within one week. In this group, 3 rats died of respiratory distress due to overdose anesthesia and one rat died during the 30 day-observation. The blood glucose in the isoflurane group was mildly increased after operation, while the 24-hour food intake and blood glucose did not markedly changed, the body mass was stably increased, and no rat died during the 30-day-observation. Conclusions Intraperitoneal injection of chloral hydrate is not suitable for intraventricular catheterization in rats. Intraperitoneal injection of pentobarbital sodium can be only carefully applied for intraventricular catheterization under poorly-limited conditions. Isoflurane inhalation anesthesia is recommended for intraventricular catheterization in rats.

Objective To assess the alterations of negative functional connectivity(FC),its relationship with clinical symptoms,and its potential value in schizophrenia(SZ).Methods Resting-state functional magnetic resonance imaging(rs-fMRI)data were acquired from patients with SZ and healthy controls(HC).For each participant,the whole brain image was first divided into 272 regions and then the FC between each pair of these regions was calculated using Pearson's correlation coefficient.Group-level negative FCs were identified using permutation test for each group.Each of the identified negative FCs was then compared between patients and controls to identify the altered negative FCs.Then,Spearman rank correlation was adopted to examine the relationship between the altered negative FCs and clinical variables.Finally,to evaluate the diagnostic value of negative FC in SZ,a multivariate pattern analysis(MVPA)was performed to distinguish between SZ and HC based on negative FCs.Results Ninety-one patients with SZ and 91 HC were included in this study,and 207 negative FCs in total were identified.Among the identified 207 negative FCs,12(constituting 5.80％of the total 207 negative FCs)were significantly altered in SZ compared with HC(Bonferroni correction,P<0.05),of which 11 were significantly decreased(i.e.,closer to 0)in SZ.The correlation analyses identified 2 significant associations-one was between a negative FC and the total score of the psychotic symptoms rating scales-auditory hallucinations(r=-0.24,P=0.02)and the other was between a negative FC and the weighted total score of the scale for the assessment of thought,language,and communication(r=0.26,P=0.01).Furthermore,the model for distinguishing between SZ and HC based on negative FCs achieved a classification accuracy of 72.6％that was significantly higher than chance-level accuracy(permutation test,P<0.001).Conclusion Negative FCs are altered in patients with SZ.Given that negative FCs are associated with clinical symptoms,thus they may serve as an imaging biomarker for assisting the diagnosis of SZ.

Objective:To investigate the diagnostic accuracy of serological indicators and evaluate the diagnostic value of a new established combined serological model on identifying the minimal hepatic encephalopathy (MHE) in patients with compensated cirrhosis.Methods:This prospective multicenter study enrolled 263 compensated cirrhotic patients from 23 hospitals in 15 provinces, autonomous regions and municipalities of China between October 2021 and August 2022. Clinical data and laboratory test results were collected, and the model for end-stage liver disease (MELD) score was calculated. Ammonia level was corrected to the upper limit of normal (AMM-ULN) by the baseline blood ammonia measurements/upper limit of the normal reference value. MHE was diagnosed by combined abnormal number connection test-A and abnormal digit symbol test as suggested by Guidelines on the management of hepatic encephalopathy in cirrhosis. The patients were randomly divided (7∶3) into training set ( n=185) and validation set ( n=78) based on caret package of R language. Logistic regression was used to establish a combined model of MHE diagnosis. The diagnostic performance was evaluated by the area under the curve (AUC) of receiver operating characteristic curve, Hosmer-Lemeshow test and calibration curve. The internal verification was carried out by the Bootstrap method ( n=200). AUC comparisons were achieved using the Delong test. Results:In the training set, prevalence of MHE was 37.8% (70/185). There were statistically significant differences in AMM-ULN, albumin, platelet, alkaline phosphatase, international normalized ratio, MELD score and education between non-MHE group and MHE group (all P<0.05). Multivariate Logistic regression analysis showed that AMM-ULN [odds ratio ( OR)=1.78, 95% confidence interval ( CI) 1.05-3.14, P=0.038] and MELD score ( OR=1.11, 95% CI 1.04-1.20, P=0.002) were independent risk factors for MHE, and the AUC for predicting MHE were 0.663, 0.625, respectively. Compared with the use of blood AMM-ULN and MELD score alone, the AUC of the combined model of AMM-ULN, MELD score and education exhibited better predictive performance in determining the presence of MHE was 0.755, the specificity and sensitivity was 85.2% and 55.7%, respectively. Hosmer-Lemeshow test and calibration curve showed that the model had good calibration ( P=0.733). The AUC for internal validation of the combined model for diagnosing MHE was 0.752. In the validation set, the AUC of the combined model for diagnosing MHE was 0.794, and Hosmer-Lemeshow test showed good calibration ( P=0.841). Conclusion:Use of the combined model including AMM-ULN, MELD score and education could improve the predictive efficiency of MHE among patients with compensated cirrhosis.

OBJECTIVETo observe the improvement of acupuncture and moxibustion on symptoms of-deficiency syndrome as well as their differences on the parameters of heart rate variability (HRV).

METHODSThirty patients with-deficiency syndrome and 15 healthy volunteers were recruited. Thirty patients with-deficiency syndrome were randomly assigned into an acupuncture group and a moxibustion group, 15 cases in each one. Fifteen healthy volunteers were allocated as a healthy control group. Patients in the acupuncture group and healthy control group were treated with acupuncture while patients in the moxibustion group were treated with moxibustion. Guanyuan (CV 4) and Zusanli (ST 36) were chosen for treatment, once every other day, for totally 10 times. All the patients were evaluated with-deficiency assessment scale (QDAS) and HRV parameters before treatment, after 4th treatment and after all treatment. The correlation was analyzed between QDAS and HRV parameters, and HRV parameters were compared among the three groups before treatment, after 4th treatment and after all treatment.

RESULTSCompared before treatment, the scores of QDAS were decreased in the acupuncture group and the moxibustion group after 4th treatment and after all treatment (all<0.05); after all treatment the score of QDAS in the moxibustion group was lower than that in the acupuncture group (<0.05). The HRV parameters of-deficiency syndrome were significantly lower than those of healthy volunteers with higher correlation with QDAS. Compared before treatment, the mean heart rate was decreased after treatment (<0.05), while total HRV and low frequency were increased in the moxibustion group (both<0.05). The mean heart rate in the healthy control group was increased after treatment (<0.05). The differences of HRV parameters before and after treatment were not significant in the acupuncture group (all>0.05).

CONCLUSIONSTotal HRV can reflect the severity of-deficiency syndrome. Both acupuncture and moxibustion can improve symptoms of-deficiency patients, which is superior in moxibustion. The possible mechanism is likely to be related with improved sustainable activation of autonomic nervous system.