Comparison of two techniques for quantifying environmental contaminants in human serum

Peak area matching and linear regression were used to quantify eight chlorinated pesticides and polychlorinated biphenyls (as Aroclor 1260) in human serum. There are no statistically significant differences in data obtained by these two quantifying techniques which were indicated by the paired t-test. For chlorinated pesticides, p = 0.053-0.62, and for polychlorinated biphenyls (as Aroclor 1260), p = 0.64. Analyte residues for the chlorinated pesticides ranged from 0.5 ppb for hexachlorobenzene (HCB) to 186 ppb for dichlorodiphenyldichloroethylene (DDE). Analyte residues for the polychlorinated biphenyls (as Aroclor 1260) ranged from 5-114 ppb. The absolute mean percent difference between the two quantifying techniques ranged from 0.06% for DDE to 8.06% for dieldrin (HEOD) among the chlorinated pesticides. The absolute mean percent difference between the two quantifying techniques for the polychlorinated biphenyls (as Aroclor 1260) was 3.4%. Peak area matching and linear regression were found to be comparable for quantifying these environmental residues in serum when the following conditions apply: 1) the concentration of the chlorinated pesticides is greater than or equal to 0.5 ppb (e.g., HCB, hexachlorocyclohexane (HCCH), oxychlordane (OC), heptachlor epoxide (HE), transnonachlor (TN), HEOD, and dichlorodiphenyltrichloroethane (DDT); 2) the concentration of the chlorinated pesticide is greater than or equal to 3 ppb (e.g., DDE); and 3) the total concentration of polychlorinated biphenyls (e.g., as Aroclor 1260) is greater than or equal to 5 ppb.     

Type I Antifreeze Proteins Enhance Ice Nucleation above Certain Concentrations

In this study, we examined the effects that antifreeze proteins have on the supercooling and ice-nucleating abilities of aqueous solutions. Very little information on such nucleation currently exists. Using an automated lag time apparatus and a new analysis, we show several dilution series of Type I antifreeze proteins. Our results indicate that, above a concentration of ∼8 mg/ml, ice nucleation is enhanced rather than hindered. We discuss this unexpected result and present a new hypothesis outlining three components of polar fish blood that we believe affect its solution properties in certain situations.     

Physiological adaptations of an under-ice population of Pseudocalanus in Barrow Strait (N.W.T) to increasing food supply in spring

Summary Zooplankton and water samples were collected at weekly intervals between April 25 and May 30, 1986 in Barrow Strait, N.W.T. (Canadian Arctic Archipelago). In tows from 0–30 m, the zooplankton community (>202 m) was dominated by Pseudocalanus. The population was apparently growing and developing as shown by an increase in the proportion of adults (stage VI) and decreases in the proportion of stages III, IV, and V as the season progressed. Respiration and excretion rates of the Pseudocalanus populations were probably linked, there being an immediate increase in excretion rate, accompanying an increase in feeding rate when chlorophyll concentrations increased, which was followed by a smaller increase in respiration rate after a time lag. Hence, there was a large decrease in the ON ratio. Increased metabolism coincided with changes in the population structure, as did protease and bodily protein, but could not be clearly linked to dietary acclimation. Only laminarinase activity could be statistically related to an identifiable fraction of potential nutritional value in the water, particulate soluble carbohydrate, but neither showed overall seasonal change.     

Ageing in the chick lens: in vitro studies.

In principle, ageing may be due to the interaction of several factors, including the accumulation of random changes both genomic and non-genomic, secondary changes in a tissue contingent upon the changing function of other tissues, and programmed non-random changes in the tissue-specific expression of various genes. The use of a single tissue comprising one cell type only, in which the major gene products are well defined, in which there is a well attested series of developmental and age-related changes in cell properties and gene expression and which can be studied and compared in vivo and in vitro, offers advantages for investigation of these questions. The vertebrate eye lens possesses these advantages. The crystallins (proteins expressed at super-abundant levels in the lens) are well characterised. The lens epithelial cells (LEC) grow readily and can differentiate into the lens fibre cells in vitro, and, finally, such terminally differentiated cells may also be derived, by a process of transdifferentiation, from neural retina cells (NRC) in vitro. Thus the effect on ageing changes of the tissue of origin may also be studied. This article reviews our previous studies on long-term changes in growth potential, differentiation capacity and crystallin expression of chick lens cells in ageing cultures, their overall similarity to events in vivo and the effect on ageing changes of genotypes affecting the growth rate. It presents new information on these genetic aspects, and on crystallin expression in long-term ageing cultures of transdifferentiated neural retina, and compares the behaviour of ageing chick lens cells with that reported for mammals.     

The majority of independently transformed BHK cell clones share a single functional lesion which determines anchorage independence and influences tumorigenicity

Summary Expression of the anchorage-independent transformed phenotype in BHK 21/13 cells generally behaves as a recessive trait. When chemically induced and spontaneously arising transformants are fused to the nontransformed parent line, transformation is initially suppressed, reappearing after extended growth of the hybrids. In this paper, complementation for the expression of anchorage independence was sought among a large group of such transformants, all independently derived from BHK 21/13 cells. Tumorigenicity studies on selected hybrids and parental lines indicated that the in vitro trait of anchorage independence is an accurate indicator of in vivo neoplasia for these cells. Seventeen of the 18 clones tested did not complement one or more of three tester strains. This result indicates that anchorage independence arose in these clones as a result of lesions in the same genetic function and suggests that the final step in the progressive changes of carcinogenesis may frequently be restricted to lesions at a single locus. This investigation was supported by National Institutes of Health grant CA27306.     

Immunoassay of muscle-specific creatine kinase with a monoclonal antibody and application to myogenesis and muscular dystrophy.

A competition e.l.i.s.a. (enzyme-linked immunosorbent assay) is described that enables direct measurement of the muscle-specific polypeptide of chick creatine kinase (M-CK) in extracts of differentiating muscle-cell cultures and in blood plasma samples, even in the presence of embryonic, or brain-type, creatine kinase. The characteristics of the assay can be considerably improved by the use of a monoclonal antibody, CK-ART, instead of rabbit antisera, and we offer an explanation for this in terms of heterogeneity of antibody affinities in polyclonal antisera. In addition to native enzyme, the assay will measure creatine kinase unfolded and inactivated by 8 M-urea treatment. During chick muscle differentiation in vitro, M-CK increased from 7.5% of the total creatine kinase at 24h to 76.0% at 143h, in good agreement with isoenzyme separation data. As a percentage of the total cell protein, M-CK increased by 156-340-fold over the same period and constituted 0.38-0.56% of the total protein in late cultures. E.l.i.s.a. measurements on 17-20-day embryonic thigh-muscle extracts, which contain almost exclusively M-CK, agree well with enzyme activity and radioimmunoassay. M-CK constituted 0.7-1.6% of the total protein in 17-19-day embryonic thigh muscle. Plasma M-CK concentrations in normal 2-8-week-old chickens were found to be in the range 0.5-0.9 micrograms/ml. Plasma concentrations of 32-56 micrograms/ml were found in 8-week-old dystrophic chickens by both e.l.i.s.a. and enzyme-activity measurements. The results suggest that inactive or unfolded forms of M-CK do not normally exist, in any significant amounts, in cell and tissue extracts or in freshly prepared samples of plasma.     

Two low-molecular-weight Ca2+-binding proteins isolated from squid optic lobe by phenothiazine--Sepharose affinity chromatography.

We have isolated two Ca2+-binding proteins from squid optic lobes, each of which is also able to bind phenothiazines in a Ca2+-dependent manner. These proteins have each been purified and partly characterized. One of the proteins corresponds to calmodulin, in that it has a similar amino acid content to bovine brain calmodulin, including a single residue of trimethyl-lysine, it co-migrates with bovine calmodulin both on alkaline-urea- and on sodium dodecyl sulphate (SDS)/polyacrylamide-gel electrophoresis, and will activate calmodulin-dependent phosphodiesterase. The second protein has the same subunit molecular weight as calmodulin, as determined by SDS/polyacrylamide-gel electrophoresis, Mr 17 000, but migrates more slowly than this protein on alkaline-urea-gel electrophoresis. It has an amino acid composition distinct from calmodulin, containing no trimethyl-lysine, its CNBr fragments migrate on alkaline gels in a pattern distinct from those of calmodulin and it shows little ability to activate phosphodiesterase. The u.v.-absorption spectra of the proteins indicate the absence of tryptophan and the presence of a high phenylalanine/tyrosine ratio in each. Both proteins also bind 3-4 calcium ions/mol at 0.1 mM-free Ca2+ and each binds chlorpromazine in a Ca2+-dependent manner.     

Solubilization, characterization, and detergent interactions of lymphocyte 5'-nucleotidase

5'-Nucleotidase is a member of a recently identified class of membrane proteins that is anchored via a phosphatidylinositol-containing glycolipid. The enzyme was readily solubilized with full retention of catalytic activity by nonionic and anionic detergents such as alkylthioglucosides, deoxycholate, and 3-[(3-cholamidopropyl)-dimethylammonio]-1-propane-sulfonate (CHAPS), while the cationic detergent dodecyltrimethylammonium bromide (DTAB) caused loss of activity. 5'-Nucleotidase was released only at high detergent concentrations, suggesting that it is tightly associated with the membrane. DTAB and deoxycholate caused a loss of heat stability, while alkylthioglucosides had no effect. CHAPS produced a remarkable increase in the heat stability of the partially purified (glycoprotein fraction) and purified enzyme. Arrhenius plots of solubilized 5'-nucleotidase showed "break points" for all detergents in the temperature range 30-37 degrees C. SDS-PAGE of pure 5'-nucleotidase showed a single subunit of molecular mass 70 kilodaltons (kDa), while sucrose density gradient sedimentation gave a peak of activity corresponding to 132 kDa, indicating that the enzyme exists as a dimer. Gel filtration of the solubilized enzyme in several detergents showed apparent molecular masses between 200-630 kDa, suggesting that lymphocyte 5'-nucleotidase may be present in high molecular mass aggregates in its native state.     

Coordinate and independent regulation of αB-crystallin and HSP27 expression in response to physiological stress

α-Crystallins share structural and functional properties with the stress protein hsp27. These polypeptides are expressed at low constitutive levels in many tissues including brain, and αB-crystallin and hsp27 can accumulate in central nervous system glia in a variety of neurological conditions. We report here that heat shock and exposure to transition metals result in an increase in the steady state mRNA level of αB-crystallin and hsp27 in primary cultures of rat forebrain astrocytes. Both exposure to tumour necrosis factor-α and hypertonic conditions result in αB-crystallin mRNA accumulation but no change in the hsp27 mRNA level. Under some of these conditions increased synthesis and accumulation of αB-crystallin and hsp27 protein are also evident. We are unable to detect αA-crystallin mRNA in resting or stressed astrocytes. A novel phenomenon involving a transitory change in stress protein mRNA mobility in Northern blots during induction is reported, which is stress type and cell type independent. The results demonstrate multiple stress regulation of αB-crystallin and hsp27 in cultured astrocytes, suggesting that they can legitimately be regarded as stress proteins in the central nervous system. © 1994 wiley-Liss, Inc.     

Exploring the legacy of African and Indigenous Caribbean admixture in Puerto Rico

Objectives

From an anthropological genetic perspective, little is known about the ethnogenesis of African descendants in Puerto Rico. Furthermore, historical interactions between Indigenous Caribbean and African descendant peoples that may be reflected in the ancestry of contemporary populations are understudied. Given this dearth of genetic research and the precedence for Afro-Indigenous interactions documented by historical, archeological, and other lines of evidence, we sought to assess the biogeographic origins of African descendant Puerto Ricans and to query the potential for Indigenous ancestry within this community.

Materials and Methods

Saliva samples were collected from 58 self-identified African descendant Puerto Ricans residing in Puerto Rico. We sequenced whole mitochondrial genomes and genotyped Y chromosome haplogroups for each male individual (n = 25). Summary statistics, comparative analyses, and network analysis were used to assess diversity and variation in haplogroup distribution between the sample and comparative populations.

Results

As indicated by mitochondrial haplogroups, 66% had African, 5% had European, and 29% had Indigenous American matrilines. Along the Y chromosome, 52% had African, 28% had Western European, 16% had Eurasian, and, notably, 4% had Indigenous American patrilines. Both mitochondrial and Y chromosome haplogroup frequencies were significantly different from several comparative populations.

Discussion

Biogeographic origins are consistent with historical accounts of African, Indigenous American, and European ancestry. However, this first report of Indigenous American paternal ancestry in Puerto Rico suggests distinctive features within African descendant communities on the island. Future studies expanding sampling and incorporating higher resolution genetic markers are necessary to more fully understand African descendant history in Puerto Rico.