Developmental comparison between cerebral organoids in vitro and body's cortices in vivo / 解剖学报

Objective To understand the characteristics and developmental differences between cerebral organoids in vitro and normal cerebral cortices in vivo. Methods 1. Grouping: cerebral cortices in vivo group and cultured cerebral organoids in vitro group. 2. Sample collection: cortical tissues were collected from Kunming mouse embryos at embryonic day 7.5(E7.5), E9.5, E11.5, E14.5, and postnatal day 3 (P3) or P7. Three specimens were taken from each group. Meanwhile, cerebral organoids were cultured with mouse induced pluripotent stem cells (iPSCs), and samples at different culture time point were collected, and more than 3 samples were collected at each time point. 3. Detection method: the distribution of different types of cells in each group of specimens was analyzed by immunofluorescent staining. Results While relative similarities between in vivo cerebral cortical development and the cerebral organoids in vitro were observed, including the histogenesis, and the morphological differentiation of nerve cells and glial cells, the lamellar architecture of cerebral cortex in mouse brain was not observed in cerebral organoids. Conclusion The development of cerebral organoids in vitro has some similarity with body's cortical development. Therefore, cerebral organoids can be used to a substitution of cortex and diseases' models, but improvement of the existing technologies is necessary.

Role and mechanism of protein kinase D in the replication of human coronavirus 229E / 中国人兽共患病学报

This study investigated the role of protein kinase D(PKD)in the replication of human coronavirus 229E(HCoV-229E)and its potential molecular mechanisms.Using MRC-5 cells as a model,we assessed the expression levels of PKD family genes and proteins through qPCR and western blotting.We knocked down Prkd3 with siRNA and inhibited PKD activity with CRT0066101 to examine the replication lev-els of HCoV-229E-infected cells.Cell viability was assessed with CCK8,viral titration was determined with the TCID50,and immunofluorescence staining was used to assess HCoV-229E expression and the structure of the trans-Golgi network(TGN).The level of phosphatidylinositol 4,5-bisphosphate(PI(4,5)P2)was measured to reflect phosphatidylinositol 4-kinaseⅢβ(PI4KⅢβ)activity.PI4KⅢβ activity was inhibited with a PI4KⅢβ inhibitor(BQR695)to observe changes in TGN struc-ture and HCoV-229E replication after cell infection.CRT0066101 was used to inhibit PKD activity in Vero-E6 cells,and HCoV-229E replication levels were examined post-infection.The expression levels of PKD family genes and proteins,including Prkd1,Prkd2 and Prkd3,significantly increased during HCoV-229E infection.Knockdown of PKD3 significantly inhibited HCoV-229E mRNA levels and titers with respect to those in the control group(t=8.999,P＜0.001;t=6.920,P＜0.001),whereas overexpression of PKD3 increased HCoV-229E mRNA levels and titers(t=6.630,P＜0.001;t=5.794,P＜0.001).CRT0066101 also significantly inhibited HCoV-229E mRNA levels and titers(t=6.931,P＜0.001;t=4.055,P＜0.01).Im-munofluorescence staining indicated that CRT0066101 inhibited TGN fission caused by HCoV-229E infection.Inhibition of PI4KⅢβ activity significantly decreased PI(4,5)P2 levels,and BQR695 rescued the elevated PI(4,5)P2 levels caused by PKD overexpression(t=6.671,P＜0.01).BQR695 decreased TGN fission in HCoV-229E-infected cells,and HCoV-229E mRNA levels and titers(t=5.151,P＜0.001;t=7.744,P＜0.001).CRT0066101 inhibited HCoV-229E mRNA levels and titers in Vero-E6 cells(t=7.480,P＜0.001;t=7.228,P＜0.01).This study revealed that PKD regulates TGN fission through modu-latingPI4KⅢβ and is involved in HCoV-229E replication.PKD inhibitors and PI4KⅢβ inhibitors significantly decreased HCoV-229E replication,thus providing important insights for future research onthe treatment of coronavirus infections.

Serum Metabolomic Indicates Potential Biomarkers and Metabolic Pathways of Pediatric Kashin-Beck Disease / 生物医学与环境科学（英文）

Objective@#To explore potential serum biomarkers of children with Kashin-Beck Disease (KBD) and the metabolic pathways to which the biomarkers belong.@*Methods@#A two-stage metabolomic study was employed. The discovery cohort included 56 patients, 51 internal controls, and 50 external controls. The metabolites were determined by HPLC-(Q-TOF)-MS and confirmed by Human Metabolome Databases (HMDB) and Metlin databases. MetaboAnalyst 3.0 and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database were used to analyze the metabolic pathways of the candidate metabolites. The use of HPLC-(Q-TRAP)-MS enabled quantitative detection of the target metabolites which were chosen using the discovery study and verified in another independent verification cohort of 31 patients, 41 internal controls, and 50 external controls.@*Results@#Eight candidate metabolites were identified out in the discovery study, namely kynurenic acid, N-α-acetylarginine, 6-hydroxymelatonin, sphinganine, ceramide, sphingosine-1P, spermidine, and glycine. These metabolites exist in sphingolipid, glutathione, and tryptophan metabolic pathways. In the second-stage study, five candidate metabolites were validated, including kynurenic acid, N-α-acetylarginine, sphinganine, spermidine, and sphingosine-1P. Except for spermidine, all substances exhibited low expression in the case group compared with the external control group, and the difference in levels of sphinganine, spermidine, and sphingosine-1P was statistically significant.@*Conclusion@#The direction of change of levels of sphinganine, spermidine, and sphingosine-1P in the two-stage study cohorts was completely consistent, and the differences were statistically significant. Therefore, these substances can be used as potential biomarkers of KBD. Furthermore, these results raise the possibility that sphingolipid metabolic pathways may be closely related to KBD.

Serum Metabonomics of Articular Cartilage Destruction Induced by T-2 Toxin in Wistar Rats / 生物医学与环境科学（英文）

The molecular pathogenesis of T-2 toxin-induced cartilage destruction has not been fully unraveled yet. The aim of this study was to detect changes in serum metabolites in a rat anomaly model with articular cartilage destruction. Thirty healthy male Wistar rats were fed a diet containing T-2 toxin (300 ng/kg chow) for 3 months. Histopathological changes in femorotibial cartilage were characterized in terms of chondrocyte degeneration/necrosis and superficial cartilage defect, and the endogenous metabolite profile of serum was determined by UPLC/Q-TOF MS. Treated rats showed extensive areas of chondrocyte necrosis and superficial cartilage defect in the articular cartilage. In addition, 8 metabolites were found to change significantly in these rats compared to the control group, including lysoPE (18:0/0:0), lysoPC(14:0), lysoPC[18:4 (6Z,9Z,12Z,15Z)], lysoPC[(16:1(9Z)], lysoPC(16:0), L-valine, hippuric acid, and asparaginyl-glycine. These 8 metabolites associated with cartilage injury are mainly involved in phospholipid and amino acid metabolic pathways.

Differential expression profiling of A549 cells induced by Toxoplasma polymorphism effector ROP16 / 中国人兽共患病学报

The aim of this study is to explore the changes and significance of gene expression profile induced by ROPl6 of Toxoplasma gondii (T.gondii)type Ⅰ,Ⅱ,Ⅲ.The A549 cell line was used as the research model.And,stable transfected ROPl6 of type Ⅰ,Ⅱ,Ⅲ of T.gondii cell lines were established.Then,we used gene expression microarray to identify genes that were differentially expressed.Gene Ontology (GO)functional classification and pathway analysis were performed on screened differentially expressed genes.According to the results of the gene expression profile,ROP16Itransfer group owned 1 270 different genes,with 776 genes up-regulated and 494 genes down-regulated;ROP16Ⅲtransfer group owned 1 362 differ-ent genes,with 842 genes up-regulated and 520 genes down-regulated;ROP16Ⅱtransfer group owned 11 052 different genes, with 5 063 genes up-regulated and 5 989 genes down-regulated.The ROP16Ⅰ/Ⅲgroup has the similar expression spectrum. However,ROP16Ⅱtransfer group was very different,which has distinct molecular profiles.Pathway analysis showed 74 path-way changes of the ROP16Ⅱtransfer group,and 19 pathways was solely dominated by it.In addition,these pathways were re-lated to NF-kappa B,Toll-like receptor,Chemokine signa-ling pathway and inflammatory response,immune response and metabolic process.Above all,ROP1 6 could affect host cell gene expression profiles.And,compared to Toxoplas-ma gondii type Ⅰ,Ⅲ,type Ⅱ has unique gene expression profiles and signaling pathway.Understanding its unique signaling pathway is very important for the prevention of type Ⅱ T.gondii.

Detection of the Urinary Biomarkers PYD, CTX-II, and DPD in Patients with Kashin-Beck Disease in the Qinghai Province of China / 生物医学与环境科学（英文）

Kashin-Beck disease (KBD) is an endemic degenerative osteoarthropathy of uncertain etiology. The aim of our study was to identify changes in C-telopeptide of type II collagen (CTX-II), pyridinoline (PYD), and deoxypyridinoline (DPD) among KBD patients. 54 KBD patients and 78 healthy controls were included this study. Urinary samples were collected and measured by ELISA. The median quantities of PYD, CTX-II, and DPD of KBD patients were 1107.73 ng/μmol.cre, 695.11 ng/μmol.cre, and 1342.34 pml/μmol.cre, while the median quantities of healthy controls were 805.59 ng/μmol.cre, 546.47 ng/μmol.cre, and 718.15 pml/μmol.cre, respectively. The differences between KBD patients and healthy controls were statistically significant (Z = 4.405, 3.653, and 3.724; P < 0.001). The higher levels of PYD, CTX-II, and DPD detected in KBD patients indicate that they could be used as biomarkers of KBD.

A Primary Investigation on Serum CTX-II Changes in Patients Infected with Brucellosis in Qinghai Plateau, China / 生物医学与环境科学（英文）

Brucellosis is one of the most widespread zoonotic diseases, with the most frequent complication being osteoarticular changes. The aim of this study was to assess the changes of C-terminal telopeptide of type II collagen (CTX-II) in patients infected with brucellosis. A total of 84 brucellosis patients and 43 volunteers were selected and divided into brucellosis vs. control groups. Serum samples were subjected to serological tests for brucellosis, and CTX-II levels in all samples were measured simultaneously with ELISA. The results showed that serum CTX-II levels in human brucellosis were higher than those of healthy controls, without a statistically significant difference, but serum CTX-II levels in male patients were significantly higher than those of female patients (P<0.05). This finding could indicate the biological changes in the cartilage and bone in human brucellosis.

X-ray measurement of metacarpal and phalange bones in the right hand of adult patients with Kaschin-Beck disease in Qinghai Province / 中国地方病学杂志

Objective By X-ray measurement of metacarpal and phalange bones in the right hand of adult patients with Kaschin-Beck disease in Qinghai Province,to provide anatomic data and information for clinic diagnosis of the disease and application of hand surgery.Methods Fifty-four cases of adult patients with Kaschin-Beck disease,who came from Guide County and Xinghai County in Hainan Prefecture,Qinghai Province,were selected as investigation subjects,then their right hand X-ray film were taken,metacarpal and phalange bones were measured by electronic digital vernier caliper.All data were analyzed with SPSS 19.0.Results The axis length of the metacarpal bones was in the order of:the 2nd metacarpal ＞ 3rd metacarpal ＞ 4th metacarpal ＞ 5th metacarpal ＞ first metacarpal.The axis length of proximal phalange bones was in the order of:the middle finger proximal ＞ ring finger proximal ＞ index finger proximal ＞ little finger proximal ＞ proximal thumb,while the middle,distal phalanx axis length varied greatly.The length of metacarpal and phalange bones of all male was longer than that of female(all P ＜ 0.01).The width of the metacarpal bones was in the order of:the 2nd metacarpal ＞ 3rd metacarpal ＞ 5th metacarpal ＞ 4th metacarpal.The width of the 2nd-5th metacarpal of male and female was (8.57 ± 0.90),(8.25 ± 0.80),(6.84 ± 0.69),(7.70 ± 0.77)mm and (7.40 ± 0.74),(7.36 ± 0.70),(6.00 ± 0.57),(6.62 ± 0.65)mm,respectively,the differences were significant(t =5.24,4.32,4.85,5.55,all P ＜ 0.01).The matacarpal bone index of female[(8.23 ± 0.90)mm] was significantly larger than that of male[(7.61 ± 0.76)mm,t =2.73,P ＜ 0.01].Conclusion X-ray measurement of hand may provide reference information for diagnosis of Kaschin-Beck disease and determination of metacarpal and phalange length in hand surgery.

Investigation of state and influence factors of children's Kaschin-Beck disease in Xinghai county of Qinghai province in 2009 / 中国地方病学杂志

Objective To investigate the present prevalence state of children's Kaschin-Beck disease (KBD) in Xinghai county of Qinghai province,a relative active KBD area in 2009,and to investigate their nutritional selenium level of local children and the T-2 toxin contamination level in their staple food.Methods Right hand X-ray photographs of children aged 7 - 12 in Shang,Zhong and Xia villages of Tangnaihai countryside in Xinghai county were taken.X-ray diagnosis was carried out according to the Diagnostic Criteria of Kashin Beck Disease (GB 16003-1995 ).Selected samples (children's hair,drinking water and their staple food) were collected according to X-ray film taken.Selenium contents in hair,drinking water and staple food samples were measured by 2,3-diaminonaphthalene fluorescence,and T-2 toxin in staple food sample was detected with enzyme-linked immunosorbent assay(ELISA) kits.ResultsTotal X-ray detection rate of children KBD was 12.20％(31/254) and KBD positive rate of children in Xia village was up to 14.97％(22/147),Shang village was up to 9.52％(6/63),and Zhong village was up to 6.82％ (3/44).The selenium level in children's body and outer environment was very low,namely,the selenium content in hair,drinking water,wheat and flour was (0.250 ± 0.136)mg/kg,(0.156 ± 0.046)μg/L,(0.0045 ± 0.0030)mg/kg,and (0.0067 ± 0.0116)mg/kg,respectively.The T-2 toxin level was relatively high in children's staple food,which was (78.91 ± 46.17)μg/kg in wheat and (47.47 ± 46.47)μg/kg in flour.Conclusions In relative active KBD areas of Xinghai county of Qinghai province,the children's selenium nutritional level is low,and the T-2 toxin contamination level in their staple food is relatively high,which is consistent with the distribution of local children's KBD.

Analysis of iodine nutrition of vulnerable population in the high risk region of iodine deficiency disorders in Qinghai province in 2009 / 中国地方病学杂志

Objective To investigate the iodine nutrition level of the vulnerable population in the high risk region of iodine deficiency disorders for providing a scientific basis for the development of effective preventive measures in Qinghai province. Methods The non-iodized salt coverage rate ＞ 5％ of Delingha city, the coverage rate of iodized salt and the rate of qualified iodized salt ≤80％ of Jiuzhi, Wulan, Nangqian, Zaduo, Geermu,Yushu, Gande, Chengduo, and Dulan counties(cities), the median of urinary iodine ＜ 100 μg/L of Huangyuan county of 11 areas were chosen as monitoring area in 2009. Five townships(towns) were selected by their location of east, west, south, north and center in each county (city), and one school was selected in each township (town),and 60 household salt samples were collected in each school, and quantitatively determined by direct titration of iodine(GB/T 13025.7-1999). Three townships(towns) were selected in each county(city), and two schools were selected in each township(town). The content of urinary iodine of 40 children aged 8 - 10 and 20 women of childbearing age aged 18 - 40 was analyzed by As3+-Ce4+ catalytic spectrophotometry (WS/T 107-2006). ResultsThree thousand two hundred and sixty-one edible salt samples were tested. The coverage rate of iodized salt was 79.88％.The iodized salt qualification rate was 95.16％. The qualified iodized salt consumption rate was 76.02％. The noniodized salt coverage rate was 20.12％. In Nangqian, Zaduo, Yushu and Geermu counties(cities), the non-iodized salt coverage rate was 88.89％, 45.05％, 43.00％ and 12.67％, respectively. The median of urine iodine of 2536 children aged 8 - 10 was 155.8 μg/L, with ≤50 μg/L about 13.6％ (346/2536), ＞ 100 μg/L about 67.5％(1711/2536), and it was 75.1, 94.6 μg/L in Nangqian and Zaduo counties, respectively. The median of urine iodine of 665 women of childbearing age was 129.7 μg/L, with ≤50 μg/L about 22.7％(151/665), ＞ 100 μg/L about 59.2％(394/665), and it was 21.0, 54.7, 72.7 μg/L in Zaduo, Nangqian and Chengduo counties, respectively.Conclusions Insufficient intake of iodine exits in children and women of childbearing age in high risk region of iodine deficiency disorders in Qinghai province, which should be corrected as soon as possible.

Investigation report of the effect of control measures to iodine deficiency disorders in Xining in 2009 / 中国地方病学杂志

Objective To master iodine nutritional status of people after universal salt iodization in Xining that reached the stage goal of elimination iodine deficiency disorders. Methods In the 7 counties investigated of Xining in 2009, 5 towns were randomly selected in each county, and one school was randomly selected in each town, 80 children aged 8 to 10 were randomly selected in each school, and goiter were examined, urinary iodine and salt iodine were tested. Thyroid gland goiter of children was detected by thyroid palpation, children's urinary iodine was tested by As( Ⅲ )-Ce4+ catalytic spectrophotometry, and salt iodine was tested by direct titration. Results A total of 2919 children aged 8 to 10 were examined, 31 goiter was detected, goiter rate was 1.06%(31/2919).One thousand and seventy-eight urine samples were detected, urinary iodine median was 205.3 μg/L, that lower than 20 μg/L accounted for 1.9% (20/1078), lower than 50 μg/L accounted for 4.5%(48/1078). Two thousand and seventy-nine salt samples were detected, median of salt iodine was 32.80 mg/kg, the rate of non-iodized salt was 0.87%(18/2079), the coverage rate of iodized salt was 99.13%(2061/2079), the qualified rate of iodized salt was 98.64% (2033/2061), the consumption rate of qualified iodized salt was 97.79% (2033/2079). Conclusions Prevention and control of iodine deficiency disorders has achieved remarkable results in Xining city, all indicators have reached the national standard to eliminate iodine deficiency disorders.

Effect of changing grain and selenium supplementation on control of children's Kaschin-Beck disease in Qinghai province / 中国地方病学杂志

Objective To observe the effect of changing grain and selenium supplementation for 1-year on control of children's Kaschin-Beck disease in Qinghai province. Methods Epidemiology, clinical and right-hand X-ray examination were carried out on children aged 7 - 12 years in 2008. Patients were diagnosed and divided into 3 groups by village, control group from Xinjianping village in Guide county, changing grain group from Xiemalang village in Guide county and supplying salt with selenium and iodine group from Shanglujuan and Xialujuan villages in Xinghai county. One year before and after the treatment, right-hand X-ray photograph (including carpal bones)was taken and child hair samples were collected, selenium was detected by 2,3-diaminonaphthalene fluorescence spectrophotometry. Results After 1 year prevention and control, the detectable rate of X-ray in control group was raised from 4.88%(2/41) to 12.20%(5/41) , the detection rate in changing grain group was declined from 17.54%(10/57) to 5.26%(3/57), and from 13.51%(10/74) to 5.41%(4/74) in supplying salt with selenium and iodine group. In changing grain group, there were 10 patients, 7 cases were cured, 2 patients stable, 1 case progressed,no new case;in supplying salt with selenium and iodine group of 10 patients, 7 were cured, 3 patients stable, 1 new diagnosed case;in control group, 2 patients stable, 2 new diagnosed metaphysis cases, 1 new diagnosed metaphyseal case. Compared with control group, the difference was statistically significant between changing grain group and supplying salt with selenium and iodine group(x2 = 5.49,4.14, all P ＜ 0.05). After 1 year control and prevention,hair selenium contents in control group and changing grain group were increased from (107.15 ± 42.30), (125.30 ±40.30)μg/kg to (108.32 ± 35.67), (135.38 ± 65.24)μg/kg, the difference was statistically insignificant(t = 0.01,0.68, all P ＞ 0.05), and selenium contents in supplying salt with selenium and iodine group were obviously increased from (95.62 ± 43.42)μg/kg to (197.64 ± 97.08)μg/kg (t = 5.41, P ＜ 0.05). Conclusion Changing grain and supplying selenium can prevent and control children's Kaschin-Beck disease.

Analysis on surveillance outcome of Kaschin-Beck disease from national monitoring region in Xinghai county of Qinghai province from 2003 to 2008 / 中国地方病学杂志

Objective To master the prevalence trend of Kaschin-Beck disease in Xinghai county (Shanglujuan and X.ialujuan village of Tangnaihai township) from 2003 to 2008 in order to understand changes of selenium level of internal and external environments. Methods According to monitoring method on national Kaschin-Beck disease,we carried out epidemical investigation,clinical examination and X-ray photograph on school children aged 7-12 in Xinghai county,a monitoring region,and collected samples of hair and grain. The content of selenium was detected by 2,3-naphthalene fluorescence. Results From 2003 to 2008,in Shanglujuan village,the prevalence rate of clinic examination was fluctuating between 0(0/34)-17.14%(6/35); the detectable rate of X-ray examination was fluctuating between 11.11% (3/27)-20.59% (7/34),the prevalence rates of metaphysis and extremities were fluctuating between 0 (0/27)-13.21%(7/53) and 2.63% (1/38)-11.43% (4/35). In Xialujuan village,the prevalence rate of clinic examination was fluctuating between 2.94% (1/34)-13.33% (6/45); the detectable rate of X-ray examination was fluctuating between 26.67% (12/45)-43.63%(24/55),the prevalence rate of metaphysis and extremities were fluctuating between 8.33% (6/72)-26.47% (9/34) and 13.33% (6/45)-38.18% (21/55). The selenium contents in hair samples were (139.92±92.27),(134.98±63.77)μg/kg respectively in Shanglujuan and Xialujuan village in 2003; the selenium contents in grain samples were (12.90± 7.18),(14.58±9.90)μg/kg respectively in Shanglujuan and Xialujuan village in 2005. Conclusions The prevalence state of Kaschin-Beck disease in national monitoring region is rigid and pathogenetic factors of Kaschin-Beck disease are active. Selenium levels of internal and external environments are low in this region.

Investigation on Kashin-Beck disease in Xinghai County,Qinghai Province in 2006 / 中国地方病学杂志

Objective By investigating newly diagnosed Kashin-beck disease,to understand distribution range,epidemic intensity and prevalence trend of Kashin-Beck disease(KBD)in Xinghai County.Method According to monitoring method on national KBD,we carried on epidemical investigation,clinical examination and X-ray photograph on school children aged 7-13 years in 6 villages from 4 towns in Xinghai County,and hair,grain,water and soil were sampled to detect selenium content using naphthalene fluorescence.Resuit Three hundred and seventy.three school children aged 7-13 years underwent the clinical examination,the prevalence rate was 12.06%(45/373);347 school children were went on X-ray,the detection rate of X-ray was 21.61%(75/347).The selenium content was the lowest in Quanqu Village,Ziketan Town,which were(77.70±42.04),(7.44±6.93)μg/kg respectively;the selenium content Was highest in Cainaihai Village,Qushian Town,which were(103.88±58.57),(29.58±24.11)μg/kg respectively;level of in internal and external environment in 4 KBD region were all low.Conclusion These four region are new KBD region.The prevalence state of KBD is serious,pathogenetic factor of Kashin-Beck disease is very active.