Cloning and expression of kiss2 in the zebrafish and medaka

Newly discovered kisspeptin (metastin), encoded by the Kiss1/KISS1 gene, is considered as a major gatekeeper of puberty through the regulation of GnRH. In the present study, we cloned a novel kisspeptin gene (kiss2) in the zebrafish Danio rerio and the medaka Oryzias latipes, which encodes a sequence of 125 and 115 amino acids, respectively, and its core sequence (FNLNPFGLRF, F-F form) is different from the previously characterized kiss1 (YNLNSFGLRY, Y-Y form). Our in silico data mining shows kiss1 and kiss2 are highly conserved across nonmammalian vertebrate species, and we have identified two putative kisspeptins in the platypus and three forms in Xenopus. In the brain of zebrafish and medaka, in situ hybridization and laser capture microdissection coupled with real-time PCR showed kiss1 mRNA expression in the ventromedial habenula and the periventricular hypothalamic nucleus. The kiss2 mRNA expression was observed in the posterior tuberal nucleus and the periventricular hypothalamic nucleus. Quantitative real-time PCR analysis during zebrafish development showed a significant increase in zebrafish kiss1, kiss2 (P < 0.002), gnrh2, and gnrh3 (P < 0.001) mRNA levels at the start of the pubertal phase and remained high in adulthood. In sexually mature female zebrafish, Kiss2 but not Kiss1 administration significantly increased FSH-beta (2.7-fold, P < 0.05) and LH-beta (8-fold, P < 0.01) mRNA levels in the pituitary. These results suggest that the habenular Kiss1 and the hypothalamic Kiss2 are potential regulators of reproduction including puberty and that Kiss2 is the predominant regulator of gonadotropin synthesis in fish.     

Variability in CYP2C9 allele frequency: A pilot study of its predicted impact on warfarin response among healthy South and North Indians

BackgroundWide variability exists in the frequency of pharmacogenetic markers for anticoagulant response in different populations. There is insufficient data on the prevalence of these variant genotypes in the Indian population. This study aims to determine the frequency of various genotype combinations of CYP2C9*2, *3 and VKORC1-1639G>A polymorphisms in the South and North Indians.MethodsGenotyping was carried out by PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) technique in 209 North Indians (NI) and 82 South Indians (SI). Warfarin maintenance dose was predicted for all subjects based on FDA approved genotype-based dose estimates from revised COUMADIN medication guide. Fisher exact test and χ² test were applied to compare categorical data among the SI and NI groups.ResultsIn SI and NI, the allele frequency of CYP2C9*2 was 0.006 and 0.05 (significant variation; p < 0.001); of CYP2C9*3 was 0.09 and 0.11; and of VKORC1-1639A was 0.14 and 0.19 (not significant), respectively. The variation in the frequency of combined CYP2C9/ VKORC1 genotypes revealed plausible difference in warfarin response among SI and NI. Based on the FDA approved revised dosing guidelines, significantly higher percentage of NI were likely to require intermediate dose (3–4 mg/day; p = 0.015, RR = 2.16) and were also predicted to have an increased risk of bleeding episodes and over anticoagulation (p = 0.012, RR = 1.93).ConclusionsGenotype frequency of CYP2C9 and VKORC1 SNPs is variable among the two ethno-geographically distinct Indian populations. This could translate into diverse warfarin response among the Indian population.     

Molecular diagnosis of neurological disorders in India

The last decade has seen remarkable advances in sequencing the human genes. There are more genes expressed in the brain than any other organ. The knowledge regarding the genome has led to tremendous progress in molecular characterization of the genes responsible for neurological disorders. The present review covers the molecular diagnosis of Duchenne muscular dystrophy, spinal muscular atrophy, and fragile X syndrome. These are three neurologic disorders common in India for which facilities of molecular diagnosis are currently available in the country. As a result of funding by the Department of Biotechnology of the Government of India, a number of molecular diagnostic centers are being established. It is hoped that molecular diagnosis of many more neurological disorders will soon become available in India.     

Schwannoma of infratemporal fossa in a young child

Schwannomas in children are rare. A nine-year-old boy presented with swelling right side of face and proptosis for 4 months. CT scan showed a heterogenous mass in the right infratemporal fossa and extending to the posterior ethmoid. There was erosion of the bony walls of the infratemporal fossa. A biopsy through the sublabial approach was reported as schwannoma. The tumour was approached by Weber-Ferguson approach and excised in toto with complete relief of symptoms.     

An unusual orbital blow-out fracture: a case report

The complete dislocation of the orbital contents into the maxillary antrum as a result of trauma is an extremely rare event and requires immediate exploration and repositioning of the globe with reconstruction of the orbital floor. We present the case of a 10-year-old male who sustained trauma to the orbit and was initially suspected to have developed auto-evisceration. He underwent successful exploration and reconstruction of orbital floor with remarkable improvement.     

Neurotherapeutic Effects of Pueraria mirifica Extract in Early‐ and Late‐Stage Cognitive Impaired Rats

We determined the neurotherapeutic effects of Pueraria mirifica extract (PME) and pure puerarin (PU) in comparison with 17β‐estradiol (E₂) in early‐ and late‐stage cognitive impaired rats. Rats were ovariectomized (OVX), kept for 2 and 4 months to induce early‐ and late‐stage cognitive impairment, respectively, and divided into four groups that were treated daily with (i) distilled water, (ii) 100 mg/kg of PME, (iii) 7 mg/kg of PU, and (iv) 80 µg/kg of E₂ for 4 months. The estrogen deficiency symptoms of OVX rats were abrogated by treatment with E₂ or PME, but not by treatment with PU. The mRNA level of genes associated with amyloid production (App and Bace1) and hyperphosphorylated Tau (Tau4) were upregulated together with the level of impaired cognition in the 2‐ and 4‐month OVX rats. Treatment with E₂ reduced the level of cognitive impairment more than that with PME and PU, and 2‐month OVX rats were more responsive than 4‐month OVX rats. All treatments down‐regulated the Bace1 mRNA level in 2‐month OVX rats, while PU and PME also decreased the App mRNA level in 2‐ and 4‐month OVX rats, respectively. Only PU suppressed Tau4 expression in 2‐month OVX rats. Thus, PME and PU elicit neurotherapeutic effects in different pathways, and earlier treatment is optimal. Copyright © 2016 John Wiley & Sons, Ltd.     

Establishment of tyrosinase sequence database in normally pigmented Indians and Japanese for rapid determination of novel mutations

BACKGROUND: Many mutations of the tyrosinase gene have been reported in oculocutaneous albinism type I (OCA1) patient. In the future, a greater number of novel mutations will be found as the search for pathological mutations in the tyrosinase genes of OCA patients from various ethnic origins. For rapid determination in future whether an observed mutation is a polymorphism or a novel pathological one, sequence databases of the gene of various ethnic people are needed. OBJECTIVE: We established a sequence database of the tyrosinase gene of Japanese as well as Indian people. METHOD: We collected DNA from 109 Japanese and 103 Indians with normal pigmentation and analyzed their tyrosinase gene using a direct sequencing method. RESULT: The database shows an apparent difference between the two ethnic groups in polymorphisms of the tyrosinase gene namely, Q402 allele, Y192 allele and IV2+24 insT were found in the Indian population, but not in the Japanese. On the other hand, some Japanese had IV2-21 insT but none of the Indians did. The database supports the notion that the tyrosinase gene evolved and extended separately in the two ethnic groups. And the developing database confirmed that the reported mutations causing Indian and Japanese OCA were not among the polymorphisms in the database, which conversely gives genetical proof of the "genuine" pathological mutations. CONCLUSION: Eventually, the sequence database we established will contribute to demonstrating novel mutations of albinism in Indians and Japanese.