Immunology in Africa

Africa is a continent with a large burden of both infectious and non‐communicable diseases. If we are to move forward as a continent, we need to equip our growing cadre of exceptional young scientists with the skills needed to tackle the diseases endemic to this continent. For this, immunology is among the key disciplines. Africans should be empowered to study and understand the diseases that affect them, and to perform their cutting‐edge research in their country of origin. This requires a multifaceted approach, with buy‐in from funders, overseas partners and perhaps, most important of all, African governments themselves.     

Computation of retinal contour in anisomyopia

It is well documented that myopia is associated with an increase in axial length of the posterior vitreous chamber. Whether equatorial or transverse dimensions are likewise affected in myopia is relevant to further understanding of the development of ametropia. We have utilised a computing method to determine retinal contour from real eye measurements of keratometry, A-scan ultrasonography and peripheral refraction as a means of assessing the transverse dimensions of the vitreous chamber. This technique has been applied to a 21-year-old female Caucasian anisomyope with a refractive error of R −1.50/−0.50 × 130 and L −4.00/−0.50 × 160. Anisomyopia offers a special opportunity for inter-eye comparison of different degrees of myopia. The repeatability of the technique was assessed by taking 10 separate sets of the aforementioned measurements and thus generating 10 retinal contours for each eye. We conclude that this method is repeatable and is capable of demonstrating differences between anisomyopic eyes although validation against in vivo measurements is required.     

PSNAV2.0-TNFα重组质粒的构建及其在体外的表达

目的:在前期微囊化基因工程细胞制备平台的基础上,构建分泌型人肿瘤坏死因子α的真核表达载体PSNAV2.0-TNFα重组质粒,并鉴定其蛋白的体外瞬时表达,为进一步利用该基因进行微囊化细胞移植治疗和改善疾病奠定基础。方法:实验于2006-06/2007-05在解放军总医院老年医学研究所细胞生物学实验室完成。①以含有人肿瘤坏死因子αcDNA序列的质粒为模板,通过PCR扩增获得人肿瘤坏死因子α基因片段;将其定向插入真核表达载体PSNAV2.0中,获得重组质粒PSNAV2.0-TNFα。采用SalⅠ和EcoRⅠ双酶切法、PCR法及插入片段序列测定法鉴定该质粒。②利用阳离子脂质体介导法,将其转染到人胚胎肾细胞HEK-293细胞中,构建可持续分泌人肿瘤坏死因子α的基因工程细胞,采用RT-PCR法和Western blot法检测转染细胞培养上清液中人肿瘤坏死因子蛋白的体外瞬时表达。结果:①通过SalⅠ和EcoRⅠ双酶切、PCR及测序鉴定证明:在HEK-293中插入片段正确。②采用RT-PCR和Western blot法检测表明HEK-293细胞培养上清中有人肿瘤坏死因子α蛋白,Mr17000。结论:成功构建了重组质粒PSNAV2.0-TNFα真核表达载体,转染HEK-293细胞后可有效分泌人肿瘤坏死因子α蛋白,并能分泌到细胞外。     

人羊膜间充质细胞具有分化成软骨及成骨细胞的潜能

目的:人羊膜间充质细胞具有比骨髓间充质干细胞更强的扩增能力和免疫原性低等优势。建立体外适宜的诱导培养条件,观察人羊膜间充质细胞定向分化为软骨细胞和成骨细胞的能力。方法:实验于2005-09/2006-12在贵州省细胞工程重点实验室完成。①材料来源:经产妇知情同意,无菌采集健康足月分娩新生儿胎盘6份,实验经医院医学伦理委员会批准。②实验方法:采用机械法剥离羊膜组织,二步酶消化法分离收获人羊膜间充质细胞,按2.2×10~8L~(-1)密度接种,传至第1～2代用于诱导分化实验。向软骨细胞诱导分化时,人羊膜间充质细胞按3×10~8L~(-1)密度接种,诱导培养液为含体积分数0.01的胎牛血清、10 mg/L转化生长因β1、100 nmol/L地塞米松、50 mg/L抗坏血酸、1%培养基添加物。向成骨细胞诱导分化时,人羊膜间充质细胞按6×10~7L~(-1)密度接种,诱导培养液为含体积分数0.1的胎牛血清、100 nmol/L地塞米松、50 mg/L抗坏血酸、5 mmol/Lβ-甘油磷酸。③实验评估:原代细胞用流式细胞仪分析表型,免疫细胞化学染色进行波形蛋白表达鉴定。分别于体外诱导第7,14,21,28天采用免疫细胞化学法检测软骨特异性Ⅱ型胶原的表达,细胞化学法检测蛋白聚糖的表达,钙-钴法检测成骨细胞特异性碱性磷酸酶的表达,茜素红S检测钙盐沉积情况。结果:①免疫组化与表型特征:人羊膜间充质细胞高表达间充质干细胞表面标志CD29、CD44和间充质细胞标志波形蛋白。②向软骨细胞诱导分化:诱导14 d后,人羊膜间充质细胞由长梭型逐渐变为多角形,可检测到Ⅱ型胶原蛋白表达及软骨细胞特异性细胞外基质蛋白聚糖。③向成骨细胞诱导分化:诱导21 d后,可观察到人羊膜间充质细胞的胞浆内有碱性磷酸酶表达,且可见钙盐沉积。结论:人羊膜间充质细胞具有分化成软骨细胞和成骨细胞的特性,可作为骨及软骨组织工程种子细胞的新来源。     

Differential effects of propranolol on lymphocyte rosette formation and response to plant mitogens

The effects of propranolol on various lymphocyte functions were studied to gain a better understanding of the recently demonstrated suppressive effect of propranolol on rheumatoid factor production. D- and L-propranolol at a concentration of 1 X 10⁻⁴ M inhibited the formation of human EA rosettes. The inhibition occurred within one minute of adding the compounds, was reversible, and did not affect cell viability. Addition of propranolol to preformed EA rosettes failed to disaggregate them. Patching and capping of SIg by an Fab′2 anti-IgG were inhibited at 2.5 X 10⁻⁵M and above. Propranolol at 2.5 X 10⁻⁵M also inhibited lymphocyte response to phytohemagglutinin and pokeweed mitogen without evidence of cell toxicity by trypan blue staining or absolute numbers of surviving cells. Congeners of propranolol with mainly beta adrenergic blocking properties did not show inhibitory effects. The inhibitory activities of propranolol are interpreted in terms of propranolol's membrane stabilizing effects and ability to interfere with membrane receptor movement.