Abnormal hemoglobin fractions on HPLC were seen in 327 of the 2,600 cases displayed. Of this, the beta thalassemia trait was the predominant abnormality with a total of 232 cases (8.9%). There were 15(0.6%) cases of beta thalassemia major and 16 of thalassemia intermedia. The rest comprised of Hb D Punjab (13 cases; 0.5%), Elevated Hb F (25 cases; 0.9%), Hb E (seven cases including two Hb E homozygous and five Hb E heterozygous), Double heterozygous Hb E-beta thal trait (six cases), Hb Q India (five cases), Double heterozygous Hb Q India -beta thal trait (two cases), Hb S (total cases three including one Hb S homozygous; two Hb S -beta thal trait) and one case each of Hb J Meerut, Hb D-Iran and Hb Lepore trait. Detection of this abnormal hemoglobin, particularly keeping in mind a high prevalence of Hb A2, will help in prevention of more serious hemoglobinopathies including beta thalassemia major. HPLC forms a rapid and accurate tool in early detection and management of various hemoglobin disorders.

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The blood-testis barrier (BTB) is formed by tight junctions between Sertoli cells. Results of previous studies suggested that the barrier is deficient in ets variant 5 (ETV5) gene-deleted mice; therefore, microarray data were examined for changes in tight junction-associated genes. The tight junctional protein claudin 5 (CLDN5) was decreased in testes of 8-day-old Etv5(-/-) pups. The study reported herein examined the expression of CLDN5 in wild-type (WT) and Etv5(-/-) mice and evaluated its contribution to BTB function. CLDN5 protein expression was evaluated in 8-day-old WT and Etv5(-/-) and adult WT, Etv5(-/-), and W/W(v) testes by immunohistochemistry and in 8-day-old WT Sertoli cell-enriched and germ cell-enriched fractions by immunocytochemistry. Cldn5 mRNA expression was evaluated in 0- to 20-day-old and adult WT mice and in 8-day-old and adult Etv5(-/-) mice via quantitative PCR. Tracer studies were performed in adult WT, Etv5(-/-), and W/W(v) mice. The results indicate the following: 1) CLDN5 was expressed in Sertoli cells, spermatogonia, and preleptotene spermatocytes. 2) Seminiferous epithelial CLDN5 expression depended upon both the presence of germ cells and ETV5. 3) CLDN5 expression in testicular vascular endothelium and rete testis epithelium was ETV5 independent. 4) Cldn5 mRNA expression increased in the testes of juvenile mice at the time of BTB formation. 5) Testes of Etv5(-/-) and W/W(v) mice, which are both deficient in seminiferous epithelial CLDN5 expression, had biotin tracer leakage from the interstitial space into the seminiferous tubule lumen. In conclusion, CLDN5 is expressed in the seminiferous epithelium, appears to be regulated by multiple influences, and contributes to BTB function.

Mice that are ets variant gene 5 (ETV5) null (Etv5(-/-)) undergo the first wave of spermatogenesis but lose all spermatogonial stem cells (SSCs) during this time. The SSC loss in Etv5(-/-) mice begins during the neonatal period, suggesting a role for ETV5 in SSC self-renewal during this period. Herein, we show that Etv5 mRNA was present in perinatal mouse testis and that ETV5 was expressed in fetal Sertoli cells and by germ cells and Sertoli cells during the neonatal period. Transplantation of Etv5(-/-) germ cells failed to establish spermatogenesis in W/W(v) mice testes, indicating that germ cell ETV5 has a key role in establishment or self-renewal of transplanted SSCs. The SSC self-renewal is stimulated by glial cell-derived neurotrophic factor (GDNF) acting through the RET/GDNF family receptor alpha 1 (GFRA1) receptor complex in SSCs. Immunohistochemistry, quantitative PCR, and laser capture microdissection revealed decreased RET mRNA and protein expression in spermatogonia of neonatal Etv5(-/-) mice by Postnatal Days 4-8, indicating that disrupted GDNF/RET/GFRA1 signaling may occur before initial spermatogonial stem/progenitor cell decrease. Etv5(-/-) spermatogonia had reduced proliferation in vivo and in vitro. Decreased cell proliferation may cause the observed decreases in the number of type A spermatogonia (Postnatal Day 17) and daily sperm production (Postnatal Day 30) in Etv5(-/-) mice, indicating quantitative impairments in the first wave of spermatogenesis. In conclusion, ETV5 is expressed beginning in fetal Sertoli cells and can potentially have effects on neonatal Sertoli cells and germ cells. In addition, ETV5 has critical effects on neonatal spermatogonial proliferation, which may involve impaired signaling through the RET receptor.

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