转神经生长因子基因诱导神经母细胞瘤分化的研究

目的观察转染神经生长因子（nerve growth factor，NGF）基因诱导神经母细胞瘤（neuroblastoma，NB）细胞系的分化，探讨NGF在NB细胞分化中的作用。方法取本院住院NB患儿新鲜手术标本，进行原代细胞培养并分离纯化，建立细胞系，作为细胞模型。通过脂质体法介导含有NGF基因的载体质粒转染NB细胞。倒置相差显微镜观察转染前后细胞的形态学变化；MTT法及有丝分裂指数测定细胞增殖的改变。结果转染后的NB细胞表达较高水平的NGF，细胞增殖受到抑制并发生形态学改变。结论所建立的NB为可诱导型，即N型；转染NGF基因的NB细胞表达较高水平的NGF；转染NGF基因的NB细胞系表现为增殖抑制和分化。     

γ—IFN诱导神经母细胞瘤细胞分化及TrKA表达的实验研究

本实验观察不同浓度的γ－IFN对NB细胞产生增殖抑制及诱导分化作用时,TrkA MRNA表达的变化,及其与增殖抑制,分化程度的关系。首先应用,常规培养SMS－KCNR细胞;然后用三种不同浓度的γ－IFN处理这些细胞,在不同的作用时间,通过台盼蓝拒染实验判定γ－INF对NB细胞的增殖抑制作用及相差显微镜观察NB细胞形态学变化;用RNA－PCR及Southern Blot杂交法检测γ－IFN对NB细胞的Tr-kA mRNA表达的影响。结果表明：（1）1000IU／ml,2000IU/ml,γ-INF对人NB细胞的体外增殖有抑制作用。（2）γ－IFN可诱导NB细胞分化成熟及TrkA mRNA表达水平增高,其作用随时间延长,浓度增加而增强。结论：（1）γ－IFN能够抑制NB细胞增殖并诱导其分化,同时TrkA mRNA表达增加,其作用效应,呈量效依赖关系。（2）TrkA mRNA表达水平的增高可能γ－IFN体外诱导NB细胞分化逆转的分子生物学机制之一。是     

IFNγ联合TRAIL诱导神经母细胞瘤细胞凋亡的研究

目的探讨IENγ（γ干扰素）对TRAIL（肿瘤坏死因子相关凋亡诱导配体）诱导神经母细胞瘤细胞株SMS-KCNR（KCNR）细胞凋亡的影响及其发生机制。方法应用RT-PCR方法检测IFNγ作用前后KCNR细胞Caspase8的表达；应用四甲基偶氮唑蓝（MTT）比色法及流式细胞仪（FCM）检测IFNγ、TRAIL、IFNγ＋TRAIL及IFNγ＋Caspase8抑制剂（zIETD-FMK）＋TRAIL对KCNR细胞生长及凋亡的影响；应用比色法测定Caspase8相对活性。结果KCNR细胞不表达Caspase8，IFNγ作用48h后的KCNR细胞Caspase8表达明显增加；KCNR细胞对TRAIL不敏感，IFNγ诱导表达Caspase8的KCNR细胞对TRAIL敏感；IFNγ＋TRAIL组Caspase8相对活性明显高于TRAIL组及抑制剂组；zIETD-FMK能阻断Caspase8的活化而抑制TRAIL对KCNR细胞的诱导凋亡作用。结论IFNγ通过诱导Caspase8表达而逆转KCNR细胞对TRAIL诱导凋亡的耐受。     

Caspase-8在TRAIL诱导神经母细胞瘤细胞凋亡中的作用

目的：应用干扰素(IFNγ)诱导神经母细胞瘤（neuroblastoma, NB）细胞caspase 8的表达并观察其是否可以恢复NB细胞对肿瘤坏死因子相关凋亡诱导配体（TRAIL）的敏感性。方法：应用RT PCR方法检测IFNγ作用前后NB细胞caspase-8 mRNA的表达;应用Alamar Blue法及流式细胞术检测IFNγ、TRAIL、IFNγ+TRAIL、IFNγ+caspase-8抑制剂zIETD-FMK+TRAIL对NB细胞生长及凋亡的影响;应用比色法测定NB细胞caspase-8相对活性。结果：对TRAIL敏感的CHP212细胞表达caspase-8,且经IFNγ处理后caspase-8 表达水平逐步增加;对TRAIL不敏感的SY5Y细胞不表达caspase-8,IFNγ作用后其caspase-8 mRNA表达明显增加。IFNγ与TRAIL联用对SY5Y细胞有明显诱导凋亡作用。CHP212细胞caspase-8相对活性随TRAIL作用时间的延长逐步升高;IFNγ与TRAIL联合作用的SY5Y细胞caspase-8相对活性明显高于未加药物处理的对照组、IFNγ组、TRAIL组及抑制剂组。结论：表达caspase-8的NB细胞对TRAIL的诱导凋亡作用敏感,TRAIL诱导NB细胞凋亡过程中伴随caspase-8活性的增加。［中国当代儿科杂志,2010,12(11)：902-907］     

肿瘤坏死因子凋亡诱导配体联合顺铂对人横纹肌肉瘤细胞株凋亡的影响

目的探讨肿瘤坏死因子相关凋亡诱导配体（TRAIL）蛋白及TRAIL基因和顺铂联合应用对人横纹肌肉瘤细胞生长抑制和诱导凋亡作用，并分析细胞线粒体膜电位和细胞内cFLIPmRNA表达的改变对凋亡的的影响。方法将TRAIL蛋白、TRAIL基因和顺铂作用于培养的人横纹肌肉瘤细胞，通过MTT比色法、流式细胞仪（FCM）检测细胞凋亡和线粒体跨膜电位的改变、RT-PCR检测cFLIPmRNA表达，观察和分析TRAIL蛋白及TRAIL基因单独对横纹肌肉瘤细胞的作用及和顺铂联合作用的效果和机制。结果TRAIL基因和100μg／L的TRAIL蛋白对横纹肌肉瘤细胞的生长抑制率分别为52．5％和43．5％，凋亡诱导率为12．95％和10．26％，联合应用顺铂，生长抑制率和凋亡诱导率均显著高于单独应用，FCM分析显示联合应用降低了线粒体跨膜电位，RT—PCR显示cFLIPmRNA表达下降，与联合应用细胞凋亡率增加相一致。结论TRAIL蛋白和TRAIL基因能有效诱导横纹肌肉瘤细胞的凋亡从而抑制横纹肌肉瘤细胞的生长，联合应用顺铂可显著提高疗效。     

阻断TrkB-BDNF信号传导通路对神经母细胞瘤细胞生长及凋亡的影响

目的：脑源性神经生长因子(BDNF)和其酪氨酸激酶受体B（TrkB）与神经母细胞瘤(NB) 细胞的化疗耐药及恶性预后密切相关。该研究探讨阻断TrkB-BDNF信号传导通路后，NB细胞对化疗药物敏感性的变化。方法：常规培养SH-SY5Y NB细胞，用nM浓度全反式维甲酸（ATRA）诱导TrkB高表达，加入BDNF、化疗药顺铂（CDDP）和特异性酪氨酸酶抑制剂K252a处理。MTT实验方法检测应用K252a前后细胞的存活率的变化；同时应用Western-blot方法检测应用K252a前后TrkB的磷酸化水平的变化；流式细胞仪（FCM）检测细胞凋亡率；透射电镜（TEM）观察凋亡细胞的形态与结构。结果：ATRA+BDNF+CDDP组细胞的存活率及凋亡率与对照组相比，差异无显著性（P>0.05）。而经K252a处理后，细胞对顺铂的敏感性增加，细胞的存活率明显降低，凋亡率明显升高，差异有显著意义。Western-blot分析显示K252a能够阻断TrkB的磷酸化。结论：阻断TrkB-BDNF信号传导通路可提高NB的化疗敏感性，逆转耐药。     

全反式维甲酸、三氧化二砷联合地西他滨对SK-N-SH细胞系活性及分化的影响

目的通过检测全反式维甲酸(ATRA)、三氧化二砷(ATO)联合地西他滨(DAC)对神经母细胞瘤(NB)细胞系SK-N-SH活性及分化的影响,探讨上述药物联合应用提高NB疗效的可能性。方法不同浓度ATRA和DAC处理SK-N-SH细胞,测定其IC50。测定单药或联合用药处理后细胞的凋亡率并记录细胞形态;采用SPSS 20.0分析数据。结果 (1)ATRA及DAC均可抑制NB细胞增殖、促进凋亡,IC50_((ATRA))=372.5μM,IC50_((DAC))=452.5μM。(2)10μM ATRA联合DAC处理NB细胞,其凋亡率为(7.31±0.94)%,高于ATRA单药组(4.90±1.58)%和DAC单药组(4.62±0.99)%。3μM ATO联合DAC处理细胞,其凋亡率由(8.44±0.86)%增加至(15.51±1.80)%。(3)1μM、10μM浓度ATRA处理SK-N-SH细胞,开始分化的时间分别为7天和3天;随时间延长,细胞分化、死亡增多。5μM DAC单药,持续观察9天,未见分化细胞。联合用药组细胞分化出现更早。结论 (1)ATRA、DAC对SK-N-SH细胞有抑制增殖、促进凋亡和诱导分化的作用;(2)联合去甲基化药物DAC可提高ATRA对SK-N-SH细胞的杀伤作用和诱导分化作用;(3)联合去甲基化药物DAC可提高ATO对SK-N-SH细胞的杀伤作用。     

苦参碱对人神经母细胞瘤SH-SY5Y细胞的作用机制

目的 探讨苦参碱对人神经母细胞瘤( NB) SH-SY5Y细胞的作用机制.方法 取对数生长期的细胞,分为对照组(含细胞、无苦参碱)及药物处理组(苦参碱质量浓度为2.0g·L-1,对细胞的作用时间分别为16 h、24 h、32 h)共4组,每组5个复孔.应用四甲基偶氮唑蓝(MTT)比色法检测NB SH-SY5Y细胞增殖抑制率;流式细胞仪(FCM)检测细胞凋亡率;比色法检测Caspase-8的相对活性.采用SPSS 16.0统计软件进行单因素方差分析.结果 MTT检测对照组及各时间点药物处理组增殖抑制率分别为(4.98±1.20)％、(11.01±0.85)％、(15.22±0.71)％和(22.31±1.45)％;FCM法检测对照组及各时间点药物处理组细胞的凋亡率分别为(5.23±1.19)％、(10.74±1.65)％、(14.00±0.98)％和(17.81±1.06)％;比色法测定各组Caspase-8的相对活性分别为(4.25±1.00)％、(10.69±1.10)％、(14.80±1.44)％和(19.80±0.97)％;以上检测各组间比较差异均有统计学意义(Pa＜0.05).结论 苦参碱可抑制人NB SH-SY5Y细胞增殖,并可通过上调Caspase-8的活性诱导NB SH-SY5Y细胞凋亡,其作用随时间的延长逐渐增强.     

γ干扰素和神经生长因子诱导神经母细胞瘤细胞分化及TrkA的表达

目的　在人神经母细胞瘤 (neuroblastoma ,NB)SMS KCNR细胞系中分别应用γ干扰素(IFN γ)、神经生长因子 (NGF)及联合应用IFN γ和NGF作为诱导剂诱导NB细胞分化 ,观察细胞形态学的变化、细胞增殖情况及TrkAmRNA的表达。方法　常规方法培养人SMS KCNR细胞系细胞 ,收取第 10天培养的细胞 ,提取总RNA ,应用逆转录 聚合酶链反应 (RT PCR)方法检测TrkAmRNA的表达水平 ,应用锥虫兰染色方法计数活细胞数并用倒置相差显微镜观察细胞形态学的改变 ,计数细胞分化率。结果　在KCNR细胞中同时负荷IFN γ和NGF的细胞表现出比单独负荷IFN γ或NGF更加显著的形态学分化 (P <0 0 1)。前者细胞增殖抑制更加明显 (P <0 0 1)。在IFN γ组 ,TrkAmRNA表达增加 (P <0 0 1) ,NGF组TrkAmRNA的表达未见明显改变 (P >0 0 5 ) ,联合用药组TrkAmRNA的表达有明显降低 (P <0 0 1)。结论　在人SMS KCNR细胞中 ,联合应用IFN γ和NGF比单独应用二者有更加显著诱导分化的作用。与IFN γ诱导TrkAmRNA表达增加不同 ,联合用药导致TrkAmRNA表达下调 ,可能存在某种负调节机制。研究结果表明 ,联合应用IFN γ和NGF可能是治疗NB甚至是晚期NB的合理方案     

Prevalence of overweight and obesity in children and adolescents in a county in Sweden

Overweight among young people in Sweden is increasing. The aim of the present study was to investigate the frequency of overweight and obesity based on body mass index (BMI) values among children and adolescents. Overweight was defined as a BMI value > or = 91st percentile and obesity as a BMI value > 98th percentile on an international reference BMI curve. The study population included boys and girls from four age groups: 9, 12, 15 and 18 y. The data consisted of self-reported measures of height and weight that were obtained from questionnaires used in a cross-sectional study in December 1997. A validation study was performed using a part of the study population. A total of 7011 (81.7%) participants completed the questionnaire. The correlation between self-reported estimations and objective measures of height and weight was high in the oldest age groups (0.88-0.98), but lower in the 9-y-old age groups (0.37-0.72). These self-reported estimations in the 9-y-olds were excluded from further analysis. It was found that 12.3%, 11.6% and 11.4% of the boys in the 12-, 15- and 18-y-old age groups and 6.8%, 5.5% and 4.8% of the girls in the same age groups were overweight and 7.9%, 8.9% and 7.3% of the boys and 5.1%, 4.2% and 3.9% of the girls were obese. Conclusion: The prevalence of overweight and obesity was found to be high in the study population and is a serious public health problem. The prevalence of obesity was significantly higher (p < 0.05) in 15-y-old boys living in rural areas than in city and town dwellers of the same age.     

The prostate in the neonate and infant in cryptorchism and hypospadias

The prostates of 8 neonates with cryptorchism and of 4 neonates and 1 infant with hypospadias have been examined and the findings have been compared with those in 127 controls.The processes of proliferation of the prostatic tubule epithelium, squamous metaplasia and secretion were normal in 7 of the 8 cases with cryptorchism and in all 5 cases with hypospadias. These findings are in contrast to 4 other cases where cryptorchism was associated with congenital renal malformations and all of whom showed an abnormal appearance of the prostate (as described elsewhere).The secretory activity of the prostate was markedly reduced in a neonate with cryptorchism who also had extreme testicular hypoplasia and marked underdevelopment of the penis, probably indicating a prenatal hormonal imbalance.2 cases with hypospadias showed a very large and distended utricle and the possible significance of this has been discussed.Associated congenital malformations, involving other than the genital system, were present in 7 of the 8 cryptorchid cases and in all 5 cases with hypospadias, as compared with 17 of the 127 controls.     

Dietary patterns in infancy and cognitive and neuropsychological function in childhood

Background: Trials in developing countries suggest that improving young children’s diet may benefit cognitive development. Whether dietary composition influences young children’s cognition in developed countries is unclear. Although many studies have examined the relation between type of milk received in infancy and subsequent cognition, there has been no investigation of the possible effect of variations in the weaning diet. Methods: We studied 241 children aged 4 years, whose diet had been assessed at age 6 and 12 months. We measured IQ with the Wechsler Pre‐School and Primary Scale of Intelligence, visual attention, visuomotor precision, sentence repetition and verbal fluency with the Developmental Neuropsychological Assessment (NEPSY), and visual form‐constancy with the Test of Visual Perceptual Skills. Results: In sex‐adjusted analyses, children whose diet in infancy was characterised by high consumption of fruit, vegetables and home‐prepared foods (‘infant guidelines’ dietary pattern) had higher full‐scale and verbal IQ and better memory performance at age 4 years. Further adjustment for maternal education, intelligence, social class, quality of the home environment and other potential confounding factors attenuated these associations but the relations between higher ‘infant guidelines’ diet score and full‐scale and verbal IQ remained significant. For a standard deviation increase in ‘infant guidelines’ diet score at 6 or 12 months full‐scale IQ rose by .18 (95% CI .04 to .31) of a standard deviation. For a standard deviation increase in ‘infant guidelines’ diet score at 6 months verbal IQ rose by .14 (.01 to .27) of a standard deviation. There were no associations between dietary patterns in infancy and 4‐year performance on the other tests. Conclusions: These findings suggest that dietary patterns in early life may have some effect on cognitive development. It is also possible that they reflect the influence of unmeasured confounding factors.     

小儿食管裂孔疝和胃食管返流的超声检查

介绍１４０例正常小儿和７例疑为胃食管返流（ＧＥＲ）患儿的食管腹腔部分和贲门的超声显像法、形态和测量值，说明Ｂ型超声和超声多普勒双功扫描不仅有助于对食管远端和贲门部生理状态的研究，也有助于食管裂孔疝和ＧＥＲ的诊断。     

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??Up to 650 000 deaths annually are associated with respiratory diseases from seasonal influenza??indicating the high burden of influenza and its substantial social cost to the world. The incidence of Influenza is high among children. Vaccine and antiviral medicine can both prevent influenza??and health education is always neglected. Neuraminidase inhibitor remains the main antiviral drug. During the management of influenza infections??doctors should pay great attention to its complications??especially pneumonia and encephalopathy.     

Normal and abnormal height and weight development in children and adolescents in Krakow--changes and trends in the last thirty years

The aim of this study is to determine the frequency of height and weight disharmony in children and adolescents in Krakow, i.e. both overweight and underweight in relation to height and the trends in last thirty years. MATERIALS AND METHODS: 4532 individuals -- a random sample of children and adolescents (2416 boys and 2107 girls) were examined in 2000. The results of the examination were compared with data obtain in 1971 (random sample of 4090 individuals) and with results from 1983 (random sample of 6536 individuals). Percentile position of height and weight were compared: the difference of two or more percentiles channels classified the results as overweight or underweight depending on height. THE RESULTS: Of the research prove that the frequency of occurrence of dis-harmonious body built increases with age, both with regard to overweight and underweight related to height and this phenomenon is more frequent in girls. In last thirty years a progressive increase of frequency of overweight was observed, but in girls it was noticeable and statistically significant only in 1971 -- 1983.     

Resistin and leptin in breast milk and infants in early life

Background

The role of adipokines in early life is considered an emerging topic issue in nutritional researches.

Aims

To evaluate serum resistin and leptin concentrations and their relations in infants and in breast milk.

Study design

We enrolled 41 term, AGA, healthy infants, of which 23 exclusively breast-fed (BF) and 18 formula-fed (FF), aged less than 6 months. Breast milk (BM) samples were collected from 23 breastfeeding mothers of the infants enrolled. Resistin concentration in serum and BM was determined by ELISA test (Human-Resistin-ELISA, Mediagnost, Reutlingen, Germany). Leptin concentration was determined by Radioimmunoassay method (LEP-R40, Mediagnost, Reutlingen, Germany). Infants weight, length and body mass index were measured. We used Mann–Whitney test. Spearman correlation was applied. Statistical significance was set at p < 0.05. Data are reported as median and interquartile range (IR).

Results

Infants serum resistin concentration was 9.30 (5.02) ng/ml. Breast milk resistin concentration (n = 23) was 0.18 (0.44) ng/ml. Leptin concentration was 3.04 (3.68) ng/ml in infants serum and in BM was 2.34 (5.73) ng/ml. Serum resistin concentrations in BF infants correlated positively with BM resistin (r = 0.636, p = 0.035). We have shown a positive correlation between resistin and leptin in total group of infants (r = 0.44, p = 0.05), confirmed in breastfed subjects (r = 0.65, p = 0.02). No correlations were found between serum hormones and anthropometric parameters of infants.

Conclusion

Our findings show interestingly a positive correlation between resistin concentrations in BF infants serum and in BM and between resistin and leptin in infants.