Alpha-adrenoceptors in dog mesenteric vessels--subcellular distribution and number of [3H]prazosin and [3H]rauwolscine binding sites.

Binding of the alpha-adrenergic antagonists [3H]prazosin and [3H]rauwolscine to well-characterized subcellular membrane fractions isolated from dog mesenteric arteries and veins was studied. Binding of both ligands was saturable with Kd values of 0.5 +/- 0.1 nM for [3H]prazosin and 5.85 +/- 0.85 nM for [3H]rauwolscine in arteries, and 0.87 +/- 0.4 nM for [3H]prazosin and 6.6 +/- 1.5 nM for [3H]rauwolscine in veins. In veins, the maximum number of binding sites for [3H]rauwolscine was higher than that for [3H]prazosin, whereas in arteries the maximum number of binding sites for each ligand was similar. In microsomes from dog aorta, the maximum number of bindings sites for [3H]prazosin was higher than that for [3H]rauwolscine. Neuronal membrane contamination in these studies was minimized by dissection procedures and evaluated by the comparison of [3H]saxitoxin binding in various preparations. Only mesenteric veins responded functionally to agonists acting on alpha 2 adrenoceptors. This study thus identified two distinct populations of [3H]prazosin and [3H]rauwolscine binding sites in the plasma membranes of dog mesenteric vessels and suggests that a much higher density of alpha 2-compared to alpha 1-adrenoceptor binding sites is required for a contractile response.     

Compliance and its role in successful treatment of an advanced periodontal case: review of the literature and a case report.

Periodontal disease is a chronic disease that is perceived by many patients to be nonthreatening. Periodontal therapy has been shown to be less effective if a regular periodontal maintenance schedule is not followed after completion of active therapy. Periodontal maintenance is an integral part of successful periodontal therapy.     

Glutathione S-transferase isoenzymes in human lung tumors

In the present studies we have compared the levels of glutathione(GSH) and GSH-related enzymes in lung tumors and correspondingnormal tissues obtained from the same individuals. We have alsoimmunologically quantitated the relative amounts of glutathioneS-transferase (or GST-P) type antigen in tumors and adjacentnormal tissues from five patients. GST activities towards 1-chloro-2,4-dinitrobenzene (CDNB) and ethacrynic acid were found to beelevated in tumors from two out of five patients (patients #1and 4), whereas the activity towards these substrates was markedlysuppressed in the tumor tissue from one of the patients (#5).Immunotitration and Western blot studies using antibodies raisedagainst -type GST isoenzymes of human lung and placenta indicatedinduction of GST -type isoenzyme in tumors from patients #1and 4 and suppression of this isoenzyme in tumor from patient#5. The tumors from patients #2 and 3 did not show any increasein GST activity or GST -type antigen. Except for the tumor frompatient #5, the GSH content was higher in the tumors from otherpatients. GSH reductase activity was found to be elevated intumors of all the patients examined in this study. These resultsindicate that GSH and GSH related enzymes are differentiallyaltered in lung tumors and GSH levels and GST - or GST-P-typeisoenzyme(s) are not uniformly elevated in all tumors.     

Epidermal growth factor receptor regulates normal urothelial regeneration

Members of the epidermal growth factor (EGF) family and their receptors are involved in many cellular processes, including proliferation, migration, and differentiation. We have previously reported that these growth factors are expressed and have specific regulatory functions in an organ-like culture model of normal human urothelial cells. Here, we used this model to investigate the involvement of EGF receptor (EGFR) in human urothelial regeneration. Three 4-mm-diameter damaged areas were made in confluent normal human urothelial cell cultures with a biopsy punch. Regeneration was measured, on fixed stained cultures, with an image analyzer, at 4, 24, and 48 hours after injury. Cell proliferation was assessed by 5-bromo-2-deoxyuridine incorporation. To identify EGF family factors potentially involved in the healing process, we studied the effect of these factors on damaged confluent cultures and the level of expression of mRNAs extracted from these cultures. EGFR inhibition of the proliferation and migration of urothelial cells was tested with (1). a specific tyrosine kinase inhibitor (AG1478) and (2). a blocking anti-EGFR antibody (LA22). Exogenously added amphiregulin, EGF, transforming growth factor-alpha and heparin-binding EGF (HB-EGF) stimulated urothelial regeneration. The damaged areas were repaired by regrowth within 48 hours. Both AG1478 and LA22 inhibited the repair (by 50% and 30%, respectively), as well as proliferation and migration. This regeneration was accompanied by increased HB-EGF mRNA expression in cultures of cells from four of six subjects, but no corresponding change in EGFR protein level was observed. These results indicate that the EGFR signaling pathway is involved in urothelial regeneration. Our data support an autocrine role of HB-EGF in this process and suggest that the EGFR pathway is a potential therapeutic target for modulating urothelial cell proliferation.     

Novel mutations in the EXT1 gene in two consanguineous families affected with multiple hereditary exostoses (familial osteochondromatosis)

Multiple hereditary exostoses (HME) is an autosomal dominant developmental disorder exhibiting multiple osteocartilaginous bone tumors that generally arise near the ends of growing long bones. Here, we report two large consanguineous families from Pakistan, who display the typical features of HME. Affected individuals also show a previously unreported feature--bilateral overriding of single toes. Analysis using microsatellite markers for each of the known EXT loci, EXT1, EXT2, and EXT3 showed linkage to EXT1. In the first family, mutation analysis of the EXT1 gene revealed that affected individuals were heterozygous for an in-frame G-to-C transversion at the conserved splice donor site in intron 1. This mutation is predicted to disrupt splicing of the first intron and produce a frameshift that leads to a premature termination codon. In the second family, an insertion of an A in exon 8 is predicted to produce a frameshift at codon 555 followed by a premature termination, a further 10 codons downstream. In both families, an increased number of affected male subjects were observed. In affected females in family 2, phenotypic variability and incomplete penetrance were noted.     

A novel autosomal recessive non-syndromic deafness locus (DFNB35) maps to 14q24.1-14q24.3 in large consanguineous kindred from Pakistan

Autosomal recessive nonsyndromic deafness is one of the most frequent forms of inherited hearing impairment. Over 30 autosomal recessive nonsyndromic hearing loss loci have been mapped, and 15 genes have been isolated. Of the over 30 reported autosomal recessive nonsyndromic hearing loss (NSHL) loci, the typical phenotype is prelingual non-progressive severe to profound hearing loss with the exception of DFNB8, which displays postlingual onset and DFNB13, which is progressive. In this report we describe a large inbred kindred from a remote area of Pakistan, comprising six generations and segregating autosomal recessive nonsyndromic prelingual deafness. DNA samples from 24 individuals were used for genome wide screen and fine mapping. Linkage analysis indicates that in this family the NSHL locus, (DFNB35) maps to a 17.54 cM region on chromosome 14 flanked by markers D14S57 and D14S59. Examination of haplotypes reveals a region that is homozygous for 11.75 cM spanning between markers D14S588 and D14S59. A maximum two-point LOD score of 5.3 and multipoint LOD score of 7.6 was obtained at marker D14S53. The interval for DFNB35 does not overlap with the regions for DFNA9, DFNA23 or DFNB5.     

Expression of keratin K2e in cutaneous and oral lesions: association with keratinocyte activation,proliferation, and keratinization

The cytoskeleton in keratinocytes is a complex of highly homologous structural proteins derived from two families of type I and type II polypeptides. Keratin K2e is a type II polypeptide that is expressed in epidermis late in differentiation. Here we report the influence of keratinocyte activation, proliferation, and keratinization on K2e expression in samples of cutaneous and oral lesions. The normal expression of K2e in the upper spinous and granular layers of interfollicular epidermis is increased in keloid scars but showed distinct down-regulation in psoriasis and hypertrophic scars where keratinocytes are known to undergo activation. Unlike normal and psoriatic skin, K2e expression in hypertrophic and keloid scars began in the deepest suprabasal layer. In cutaneous basal and squamous cell carcinomas, K2e was absent in most tumor islands but the overlying epidermis showed strong expression. No significant K2e expression in nonkeratinized or keratinized oral epithelia, including buccal mucosa, lateral border of tongue and gingiva was detected. In oral lichen planus K2e expression was undetectable, but in benign keratoses of lingual mucosa induction of K2e along with K1 and K10 was observed. In mild-to-moderate oral dysplasia with orthokeratinization, K2e was highly expressed compared with parakeratinized areas but in severe dysplasia as well as in oral squamous cell carcinoma, K2e expression was undetectable. Taken together, the data suggest that K2e expression in skin is sensitive to keratinocyte activation but its up-regulation in oral lesions is a reflection of the degree of orthokeratinization.