Interaction of monoclonal antiparathyroid antibody with Ca2+ agonistic actions of Mn2+ in normal human parathyroid cells

Effects of the monoclonal antiparathyroid antibodies G11 and E11 on Mn2+ interaction with individual normal human parathyroid cells were studied. At 0.5mM Ca2+, 3mM Mn2+ induced a rapid transient increase in cytoplasmic Ca2+ [Ca2+i] followed by quenching of the fluorescence from the Ca2+ indicator fura-2 as Mn2+ entered into the cells. Whereas the antibody E11 had no effects, treatment with G11 abolished the Ca2+i transient and considerably delayed the entry of Mn2+. The results support the presence of a cation-sensitive receptor mechanism on parathyroid cells and indicate that the antibody G11 not only blocks the interaction between Ca2+ and this receptor mechanism but also that of Mn2+.     

Delayed-onset synergism between leukotriene B4 and prostaglandin E2 in human skin

The time-course of cutaneous inflammatory responses to LTB₄ and PGE₂ both alone and in combination has been studied in 10 healthy volunteers. LTB₄ induced a transient wheal and flare response in some subjects, maximal at 15 minutes and succeeded by an erythematous, indurated lesion at 2–4 hours. PGE₂ elicited a wheal and erythema response which resolved within 1–2 hours. Combination of LTB₄ and PGE₂ produced acute wheal and erythema responses which did not differ significantly from the summation of responses to the individual constituents of the mixture or from responses to a two-fold increase in the concentration of either component. Wheal and erythema responses persisted, however, with significant potentiation of responses 4 hours after injection. As both leukotrienes and prostaglandins are generated in acute allergic reactions, the effects of these mediators in combination could contribute to persisting and late-onset responses to allergen, in both the skin and lung. In particular, sustained responses to the combination of LTB₄ and PGE₂ might be important in the pathogenesis of inflammatory skin diseases such as psoriasis.     

Genome-Wide and Locus Specific Alterations in CDC73/HRPT2-Mutated Parathyroid Tumors

Mutations in the hyperparathyroidism type 2 (HRPT2/CDC73) gene and alterations in the parafibromin protein have been established in the majority of parathyroid carcinomas and in subsets of parathyroid adenomas. While it is known that CDC73-mutated parathyroid tumors display specific gene expression changes compared to CDC73 wild-type cases, the molecular cytogenetic profile in CDC73-mutated cases compared to unselected adenomas (with an expected very low frequency of CDC73 mutations) remains unknown. For this purpose, nine parathyroid tumors with established CDC73 gene inactivating mutations (three carcinomas, one atypical adenoma and five adenomas) were analyzed for copy number alterations and loss of heterozygosity using array-comparative genomic hybridization (a-CGH) and single nucleotide polymorphism (SNP) microarrays, respectively. Furthermore, CDC73 gene promoter methylation levels were assessed using bisulfite Pyrosequencing. The panel included seven tumors with single mutation and three with double mutations of the CDC73 gene. The carcinomas displayed copy number alterations in agreement with previous studies, whereas the CDC73-mutated adenomas did not display the same pattern of alterations at loci frequently deleted in unselected parathyroid tumors. Furthermore, gross losses of chromosomal material at 1p and 13 were significantly (p = 0.012) associated with parathyroid carcinomas as opposed to adenomas. Quantitative PCR-based copy number loss regarding CDC73 was observed in three adenomas, while all the carcinomas were diploid or showed copy number gain for CDC73 gene. Hypermethylation of the CDC73 gene promoter was not observed. Our data could suggest that CDC73-mutated parathyroid adenomas exhibit a partly unique cytogenetic profile in addition to that of carcinomas and unselected adenomas. Furthermore, CDC73-mutated carcinomas displayed losses at 1p and 13 which are not seen in CDC73-mutated adenomas, making these regions of interest for further studies regarding malignant properties in tumors from CDC73-mutated cases. However, due to the small sample size, validation of the results in a larger cohort is warranted.     

Two new Ig VH gene families in Oncorhynchus mykiss

Genes encoding the immunoglobulin heavy-chain variable region (Ig VH) in rainbow trout (Oncorhynchus mykiss) have been grouped into 11 families. While obtaining a baseline assessment of the various gene families utilized by trout in the production of secreted antibody, we discovered two new families. These proposed Ig VH families, Families XII and XIII, were rarely observed; only two VH sequence types were detected for each new family, suggesting that they may not be commonly used in response to antigens, or that the captive environment may not lead to typical exposures seen in the wild. Additionally, unlike preceding studies, we found at least one representative gene sequence for each of the 11 reported Ig VH gene families, possibly indicating that the repertoire of trout Ig VH gene families may be more universal among different stocks than previously realized. GenBank accession numbers: Family XII—DQ453185 and DQ453150; Family XIII—DQ453153 and DQ453146; others DQ453143, DQ453156, DQ831723, DQ831825.     

Delayed-onset synergism between leukotriene B4 and prostaglandin E2 in human skin

The time-course of cutaneous inflammatory responses to LTB4 and PGE2 both alone and in combination has been studied in 10 healthy volunteers. LTB4 induced a transient wheal and flare response in some subjects, maximal at 15 minutes and succeeded by an erythematous, indurated lesion at 2-4 hours. PGE2 elicited a wheal and erythema response which resolved within 1-2 hours. Combination of LTB4 and PGE2 produced acute wheal and erythema responses which did not differ significantly from the summation of responses to the individual constituents of the mixture or from responses to a two-fold increase in the concentration of either component. Wheal and erythema responses persisted, however, with significant potentiation of responses 4 hours after injection. As both leukotrienes and prostaglandins are generated in acute allergic reactions, the effects of these mediators in combination could contribute to persisting and late-onset responses to allergen, in both the skin and lung. In particular, sustained responses to the combination of LTB4 and PGE2 might be important in the pathogenesis of inflammatory skin diseases such as psoriasis.     

Comparative analysis of mycobacterial infections in wild striped bass Morone saxatilis from Chesapeake Bay

During an ongoing epizootic of mycobacteriosis, wild striped bass Morone saxatilis from Chesapeake Bay were analyzed using 3 methods for detection of either mycobacterial infection or associated granulomatous pathology. The specific detection techniques, which utilized aseptically collected splenic tissue, were histology, quantitative culture and nested PCR. Based on analysis of 118 samples, detection of infection differed significantly between the 3 methods (chi-square, p = 0.0007). Quantitative culture and nested PCR detected similar, higher rates of infection (69 and 75%, respectively) than the histological method (52%). Although primary PCR assays for a 924 to 940 bp segment of the mycobacterial 16S rRNA gene were positive for genomic DNA from mycobacterial cultures, a secondary, nested PCR reaction for an internal 300 bp gene segment was required in order to detect mycobacteria within splenic tissue. A similar rate of mycobacterial infection was present in fish collected from all sites tested. Although all detection methods found that striped bass age 4.0 to 4.9 yr had the highest positive incidence, nested PCR detected a higher frequency of mycobacterial infection in fish > or = 6.0 yr of age than the other 2 methods. Quantitative bacteriology was a more sensitive detection technique when the fish tissue contained < or = 10(3) mycobacteria g(-1).     

Parathyroid-like regulation of parathyroid-hormone-related protein release and cytoplasmic calcium in cytotrophoblast cells of human placenta.

Immunohistochemical staining of human placenta revealed intense reactivity for amino terminal and midregional parathyroid-hormone-related protein (PTHrp) in the cytotrophoblast cells and weaker staining in the syncytiotrophoblasts. The cytotrophoblasts also displayed conspicuous surface staining with the monoclonal antibodies E11 and G11, which recognize a Ca2+ receptor mechanism regulating hormone release of parathyroid cells. Cytotrophoblasts enriched on Percoll gradients or by linking surface-bound E11 to magnetic beads revealed biphasic elevation of cytoplasmic Ca2+ ([Ca2+]i) upon a stepwise rise of external Ca2+ from 0.5 to 3.0 mM, with a half-maximal effect at 1.75 mM. Individual cytotrophoblasts identified by their E11 reactivity disclosed a temporary increase of [Ca2+]i upon elevation of external Mg2+, while Mn2+ triggered both a [Ca2+]i transient and an influx of itself. These effects were efficiently blocked by the G11 antibody. Depolarization with K+ or addition of the voltage-dependent Ca2+ channel blocker verapamil had only marginal effects on [Ca2+]i. Raised extracellular calcium inhibited release of PTHrp from the cells, and this inhibition was blocked by the G11 antibody. The virtually parathyroid-identical Ca2+ regulation of [Ca2+]i may mediate feedback control of PTHrp release from the cytotrophoblasts and thereby participate in the regulation of placental Ca2+ transport.