Characterization of a highly polymorphic marker adjacent to the SLC4A1 gene and of kidney immunostaining in a family with distal renal tubular acidosis.

BACKGROUND: Mutations in the human SLC4A1 (AE1/band 3) gene are associated with hereditary spherocytic anaemia and with distal renal tubular acidosis (dRTA). The molecular diagnosis of AE1 mutations has been complicated by the absence of highly polymorphic genetic markers, and the pathogenic mechanisms of some dRTA-associated AE1 mutations remain unclear. Here, we characterized a polymorphic dinucleotide repeat close to the human AE1 gene and performed an immunocytochemical study of kidney tissue from a patient with inherited dRTA with a defined AE1 mutation. METHODS: One CA repeat region was identified in a phage P1-derived artificial chromosome (PAC) clone containing most of the human AE1 gene and the upstream flanking region. We determined its heterozygosity value in multiple populations by PCR analysis. Genotyping of one family with dominant dRTA identified the AE1 R589H mutation, and family member genotypes were compared with the CA repeat length. AE1 and vH(+)-ATPase polypeptides in kidney tissue from an AE1 R589H patient were examined by immunocytochemistry for the first time. RESULTS: This CA repeat, previously reported as D17S1183, is approximately 90 kb upstream of the AE1 gene and displayed considerable length polymorphism, with small racial differences, and a heterozygosity value of 0.56. The allele-specific length of this repeat confirmed co-segregation of the AE1 R589H mutation with the disease phenotype in a family with dominant dRTA. Immunostaining of the kidney cortex from one affected member with superimposed chronic pyelonephritis revealed vH(+)-ATPase-positive intercalated cells in which AE1 was undetectable, and proximal tubular epithelial cells with apparently enhanced apical vH(+)-ATPase staining. CONCLUSIONS: The highly polymorphic dinucleotide repeat adjacent to the human AE1 gene may be useful for future studies of disease association and haplotype analysis. Intercalated cells persist in the end-stage kidney of a patient with familial autosomal dominant dRTA associated with the AE1 R589H mutation. The absence of detectable AE1 polypeptide in those intercalated cells supports the genetic prediction that the AE1 R589H mutation indeed causes dominant dRTA.     

Localization of Hsp27 in the Rat Submandibular Gland Following the Application of Various Surgical Treatments

Salivary glands repair and regenerate following various types of injuries and surgical procedures. However, the tissue responses induced in the contralateral glands have yet to be elucidated in detail. Hsp27, a member of the heat-shock protein (Hsp) family, is strongly expressed in physiological environments, particularly during development. Hsp27 was previously shown to play a role in the regulation of acinar cell proliferation and differentiation in the rat submandibular gland.The present study performed the following surgical treatments on the right submandibular glands of adult rats: 1) duct ligation followed by unligation after one week; 2) partial sialoadenectomy; and 3) total sialoadenectomy. Immunohistochemistry for Hsp27 and Ki67 was performed in the experimental and normal contralateral glands, and localization was histologically and morphometrically analyzed.The results obtained revealed the localization of Hsp27 to the intercalated duct in the submandibular glands of non-treated rats. The expression of Hsp27 was strongly induced in both the uninjured contralateral control glands as well as treated glands of experimental rats regardless of the surgical procedure performed. The number of Hsp27-immunopositive cells increased rapidly following surgery, and subsequently returned to the same level as that in non-treated rats after 4 weeks. However, no marked changes were observed in the number of Ki67-immunopositive proliferating cells. Therefore, the change in the number of Hsp27-immunopositive cells may have contributed to compensatory hypertrophy. The results of the present study indicate that the expression of Hsp27 in the intercalated duct in the submandibular gland may play a role in the differentiation of acinar cells.     

多节段腰椎间盘突出症25例治疗体会

目的探讨椎板间小开窗治疗多节段腰椎间盘突出症的可行性。方法25例多节段腰椎间盘突出症病人,用锥板间小开窗治疗。随访时间为2~30个月,按MacNab’s评定标准。结果25例病人优15例,良8例,可2例,优良率94.4%。结论椎板间小开窗是治疗多节段腰椎间盘突出症的可行手术方法。     

风心病房颤患者右房心肌细胞闰盘结构及肌原纤维的研究

目的探讨风心病持续性房颤患者右房心肌细胞闰盘及肌原纤维的病理变化。方法选择29例风湿性二尖瓣置换术患者,根据有无持续性心房颤动分为心房颤动组(AF组)和窦律组(SR组),两组之间临床因素没有统计学差异。换瓣术中取右心耳心肌标本,电镜观察闰盘和肌原纤维。结果(1)AF组患者右房心肌闰盘缝隙裂开,缝隙中出现髓样、絮状物质,SR组患者则没有发现这种现象;(2)AF组肌原纤维的体密度为36.1±10.66%,SR组为43.72±8.9%(P<0.05),显示AF组有明显的肌原纤维溶解。结论风心病慢性AF与窦性心律患者右房肌细胞闰盘结构存在差异,AF组伴有明显的肌溶解,这些改变可能是导致风心病患者右房AF的重要病理基础。     

心房颤动患者右心房肌闰盘重构的研究

目的：探讨心房颤动（房颤）患者右心房肌细胞间闰盘相关蛋白表达变化与房颤发生并维持的意义。方法：心外科患者53例，其中房颤患者27例（房颤组），窦性心律患者26例（窦律组），在建立体外循：吓前取右心房组织。分别将心房组织提取mRNA和制作病理组织切片，部分组织送电镜检查。用RT—PCR方法检查Cx40、Cx43mRNA表达变化，用免疫组化方法检测Cx43、N-钙黏素的分布变化。结果：两组患者Cx40、Cx43mRNA表达无变化，免疫组化结果显示房颤组缝隙连接、N-钙黏素分布发生不均一，光镜、电镜证实房颤组缝隙连接发生侧边化分布明显。结论：闰盘相关蛋白表达改变在房颤的维持中发挥重要作用。     

Side-to-side linking of myocardial cells in hypertrophic cardiomyopathy: whole heart microscopic observation with tangential sections

By cross-section or longitudinal section, it is difficult to investigate longitudinal features of myocardial cells in the whole heart. Here, introducing the use of tangential sections to obtain longitudinal aspect of myocardial cells in any part of myocardium, the authors evaluated myocardium in the left ventricle in 10 normal hearts and four hearts with hypertrophic cardiomyopathy (HCM). Tangential sections were obtained by peeling the superficial layer of myocardium. After peeling the whole surface, secondary deep layer was peeled. These procedures were repeated more than five times through the wall. Intercalated discs (ICD) were observed immunohistochemically with anti-N-cadherin and antidesmoplakin. In normal hearts, myocardial cells were cut longitudinally and ran parallel in tangential sections. They linked end-to-end with simple and regular ICD with average lengths of 120-130 microm and average sarcomere numbers of 56-65. In HCM hearts, many myocardial cells were cut almost longitudinally running approximately parallel in tangential sections. Myocardial cells frequently showed side-to-side linking characterized by skewed ICD, indistinct ICD counterparts, and longitudinally arranged ICD. Two young HCM hearts had circle-shaped ICD and vacuole-like structures highlighted by immunostaining for N-cadherin, which were actually extracellular structures comparable with irregular side-to-side linking. It is considered that side-to-side linking of myocardial cells is a characteristic microscopic feature in HCM rather than myocardial disarray.     

Assessment of midcarpal deformity of the wrist using the triangulation method

PURPOSE: The purpose of this study is to present an alternative method for static radiologic assessment of the wrist for midcarpal instability (ie, palmar intercalated segmental instability [PISI] and dorsal intercalated segmental instability [DISI]). The triangulation method uses 3 anatomic landmarks observed on the standard lateral x-ray of the wrist. METHODS: A total of 125 normal lateral radiographs were measured to determine the normal range for the dorsal limb (DL) to palmar limb (PL) ratio. A 2-step process of performing triangulation is described. The first step is nonspecific screening of the radiograph and defines values greater than 1.0 as having a DISI deformity and values less than 0.5 as having a PISI deformity. The second step is used only for borderline values, which takes the position of the wrist into consideration and uses a normagram (reference chart) to match the DL:PL ratio with the radiometacarpal (RM) angle. RESULTS: The average lateral wrist position was 8.4 degrees of extension (-8.4). The average DL:PL ratio was 0.75 +/- 0.09 (range, 0.93-0.57). CONCLUSIONS: Based on these data we defined DISI deformity of the wrist as DL:PL ratios greater than 1.0, and ratios less than 0.5 representing PISI deformities. The triangulation method of assessing midcarpal alignment of the carpus is a practical and simple alternative to the traditional static radiologic method of assessing midcarpal instability of the wrist.     

Impaired fear extinction in mice lacking protease nexin‐1

The serine protease inhibitor protease‐nexin‐1 (PN‐1) has been shown to modulate N‐methyl‐d ‐aspartate receptor (NMDAR)‐mediated synaptic currents and NMDAR‐dependent long‐term potentiation of synaptic transmission. Here, we analysed the role of PN‐1 in the acquisition and extinction of classical auditory fear conditioning, two distinct forms of learning that both depend on NMDAR activity in the amygdala. Immunostaining revealed that PN‐1 is expressed throughout the amygdala, primarily in γ‐aminobutyric acid containing neurons of the central amygdala and intercalated cell masses (ITCs) and in glia. Fear extinction was severely impaired in mice lacking PN‐1 (PN‐1 KO). Consistent with a role for the basal nucleus of the amygdala in fear extinction, we found that, compared with wild‐type (WT) littermate controls, PN‐1 KO mice exhibited decreased numbers of Fos‐positive neurons in the basal nucleus after extinction. Moreover, immunoblot analysis of laser‐microdissected amygdala sub‐nuclei revealed specific extinction‐induced increases in the level of phosphorylated alpha‐calcium/calmodulin protein kinase II in the medial ITCs and in the lateral subdivision of the central amygdala in WT mice. These responses were altered in PN‐1 KO mice. Together, these data indicate that lack of extinction in PN‐1 KO mice is associated with distinct changes in neuronal activity across the circuitry of the basal and central nuclei and the ITCs, supporting a differential impact on fear extinction of these amygdala substructures. They also suggest a new role for serine protease inhibitors such as PN‐1 in modulating fear conditioning and extinction.