Endocrine and physiological responses of hatchling American kestrels (Falco sparverius) following embryonic exposure to technical short-chain chlorinated paraffins (C10-13).

Short chain chlorinated paraffins (SCCPs) are complex mixtures of polychlorinated n-alkanes, shown to bioaccumulate but with unknown effects in wild birds. The present study examined development-related effects of SCCPs on captive American kestrels (Falco sparverius) treated in ovo on embryonic day (ED) 5 by injection with technical Chloroparaffin® (C10-13, 55.5% Cl) at environmentally relevant nominal (measured) concentrations of 10 (10), 50 (29) or 100 (97) ng ΣSCCP/g egg ww, and artificially incubated until hatching (ED27-ED29). The SCCP concentrations measured in the yolk sacs of the hatchling kestrels bracketed concentrations reported in the eggs of wild birds. Uptake and deposition of these SCCPs differed between male and female hatchlings, with only males showing differences in SCCP concentrations, being highest in the high-dose males than each of the other male groups. Embryonic exposure to SCCPs suppressed glandular total thyroxine (TT4) (20-33%) and reduced circulating triiodothyronine (TT3) (37-40%) in male hatchlings only when compared to control males, but had no effect on glandular TT3 or circulating TT4 in male or female kestrels. Histological assessments of thyroid glands showed that both sexes experienced significant structural changes indicative of gland activation. These thyroid glandular changes and the variations in SCCP concentrations were related to circulating TT3 in female hatchlings. Hepatic deiodinase enzyme (D1, D2) activities were stable and no SCCP-related changes were observed in hatching success, hatchling size, or immune organ size. However, several of the thyroid function indicators were correlated with hatchling size and smaller bursas and spleens, possibly indirectly through SCCP-induced changes in thyroid function. Because changes in thyroid function were evident at concentrations measured in wild bird eggs, similar changes may occur in wild nestlings. The potential impact of these changes on thyroid-mediated growth and survival in wild birds requires further investigation.

New paradoxical three-finger toxin from the cobra Naja kaouthia venom: Isolation and characterization.

A new three-finger toxin nakoroxin was isolated from the cobra Naja kaouthia venom, and its complete amino acid sequence was established. Nakoroxin belongs to the group of "orphan" toxins, data on the biological activity of which are practically absent. Nakoroxin shows no cytotoxicity and does not inhibit the binding of α-bungarotoxin to nicotinic acetylcholine receptors of muscle and α7 types. However, it potentiates the binding of α-bungarotoxin to the acetylcholine-binding protein from Lymnaea stagnalis. This is the first toxin with such an unusual property.

Channelized ice melting in the ocean boundary layer beneath Pine Island Glacier, Antarctica.

Ice shelves play a key role in the mass balance of the Antarctic ice sheets by buttressing their seaward-flowing outlet glaciers; however, they are exposed to the underlying ocean and may weaken if ocean thermal forcing increases. An expedition to the ice shelf of the remote Pine Island Glacier, a major outlet of the West Antarctic Ice Sheet that has rapidly thinned and accelerated in recent decades, has been completed. Observations from geophysical surveys and long-term oceanographic instruments deployed down bore holes into the ocean cavity reveal a buoyancy-driven boundary layer within a basal channel that melts the channel apex by 0.06 meter per day, with near-zero melt rates along the flanks of the channel. A complex pattern of such channels is visible throughout the Pine Island Glacier shelf.

Direct cloning of a target gene from a pool of homologous sequences: complete cDNA sequence of a weak neurotoxin from cobra Naja kaouthia.

Selective cloning of the cDNA coding for a weak neurotoxin (WTX) from cobra N. kaouthia including the 5'- and 3'-non-translated regions (NTR) is described. The known amino acid sequence of WTX was used together with the nucleotide sequence of a weak neurotoxin NNAM2 from cobra Naja atra, to design WTX-specific primers for direct amplification of an internal WTX cDNA fragment by RT- PCR. The sequence of the complete WTX cDNA was determined in sequencing runs on internal PCR products, cloned 3'- and 5'-RACE-fragments and several full-length cDNA clones. The cDNA coding sequence is in excellent agreement with the previously determined WTX amino acid sequence, has a high homology with other known weak toxin cDNAs, whereas even higher homology (up to 96%) with several classes of 3-finger toxins was detected in the 59 bp 3'-NTR consensus sequence. A possible function of the highly conserved nucleotide sequence elements is discussed.

Patella height after high tibial osteotomy with internal fixation and early motion.

The purpose of this study was to compare the incidence of patella infera in patients after high tibial osteotomy treated with either postoperative immobilization or internal fixation and early range of motion. A retrospective review of 98 patients with high tibial osteotomy was done at the authors' institution. Thirty-three patients who had secondary procedures such as concomitant ligamentous reconstruction with early motion were excluded. Therefore, 69 knees in 65 patients remained in the study cohort. Group A consisted of 32 patients (34 knees) treated with postoperative immobilization, whereas Group B included 33 patients (35 knees) treated with internal fixation and early motion. The preoperative and postoperative Insall-Salvati index, Blackburne-Peel index, and angular alignment were determined for each group. Between Groups A and B, the differences in the Insall-Salvati index and the Blackburne-Peel index were statistically significant, although the difference in angular correction was not significant. With rigid fixation and early motion the Insall-Salvati index showed that there was less postoperative shortening of the patellar tendon. The relationship of the patella to the remainder of the knee was affected less adversely as evidenced by the Blackburne-Peel index. High tibial osteotomy with internal fixation and early range of motion should result in a better knee and ease the subsequent performance of a total knee arthroplasty.

Quantitation of dopamine D2 receptor mRNA in a mesencephalic cell culture using a nonradioactive competitive reverse transcription polymerase chain reaction method.

Studies on gene expression during differentiation and maturation processes have to cope with determinations of extremely low steady state levels of specific mRNA. Using the experimental model of dopamine D2 receptor (D2R) expression in a primary mesencephalic cell culture we worked out a quantitative reverse transcription polymerase chain reaction method which allows to analyze and quantify mRNA levels of cells present in a few wells of the culture. The method uses an internal cRNA standard which shares both primer binding sites and PCR product length with the target sequence. The amplicons are quantitated in microplates by hybridization with immobilized capture probes that allow for the distinction of internal standard and target sequences followed by the chemiluminescent detection of hybridized DNA. Applying this method the levels of D2 receptor mRNA of the mesencephalic cell culture on day in vitro 1 amounted to about 250 fg/microgram RNA and increased to about 1200 fg/microgram RNA on day in vitro 13-15.

Fixation of the acetabular component. The case for cement.

The long-term success rate of cemented sockets in total hip arthroplasty has been well documented in patients who are 60 years of age and older and who have had a followup of as many as 16 years. The failures with cemented sockets have been observed in young patients, patients with poor bone stock (rheumatoid and dysplastic hips) with metal backed components, and in revision surgery with loss of acetabular bone. Ranawat et al have shown that most mechanical failures of cemented socket fixation within 10 years of primary operation is attributable to failure to achieve a good fixation initially of the cement and bone. Volumetric wear of the polyethylene of a cemented socket against a 22- or 28-mm femoral head is compared with the metal backed cemented and noncemented cups. The increase in volumetric polyethylene particles with metal backed cemented sockets and noncemented sockets will induce histiocytic response. The mechanism of histiocytic invasion should be similar for cemented all polyethylene sockets and noncemented sockets. It manifests itself in the cemented socket as global radiolucency when the socket is loose and as osteolysis when it is well fixed for noncemented and cemented sockets. If the number of particles coming out in a noncemented and hybrid total hip replacement are greater, osteolysis would be expected to increase with longer followup. The technique of cemented polyethylene sockets requires organization of the surgical team and hypotensive epidural anesthesia. Under these conditions the procedure is very reproducible. As far as cost is concerned, the all polyethylene socket is significantly less expensive. It seems that cemented total hip replacement is most suitable and perhaps is the right kind of operation for treating osteoarthritis of the hip for patients who are 60 years of age and older because the procedure is reproducible, the quality of arthroplasty is excellent, and it is durable, lasting as many as 15 years in 90% to 95% of the cases.

Comparison of different quantitative PCR procedures in the analysis of the 4977-bp deletion in human mitochondrial DNA.

Four quantitative PCR procedures were compared to analysing the level of the 4977 bp deletion in human mitochondrial DNA. A single preparation of total cellular DNA from the heart of a 69-year-old female subject was used for all four methods. We estimated the deletion to represent 0.005% of total mtDNA molecules using the serial dilution procedure and two internal standard methods with two separate sets of reference recombinant plasmids. However, the value obtained with the kinetic PCR analysis was 0.5%, two orders of magnitude higher. The internal standard procedures are likely to be the most accurate among the four methods used, but the more technically convenient serial dilution method would also be an appropriate choice.

Prediction of the long-term durability of all-polyethylene cemented sockets.

A retrospective study was done to determine the correlation between the rate of aseptic loosening and the early radiographic appearance of the bone-cement interface of the acetabulum of primary cemented total hip prostheses. Two hundred thirty-six hips implanted with modern cement technique were studied during an average clinical and radiographic followup of 9 years. Early radiographs and the last available radiographs were studied by 3 independent observers, and the quality of the bone-cement interface was evaluated with a standardized scoring system. On the acetabular side, the authors noted 0.8% clinical failure, a 3% rate of radiographic migration, and a 3.4% rate of progressive global radiolucency. Only 2.2% of hips considered well fixed on the postoperative film were loose. Of the hips considered not well fixed, the rate of aseptic loosening was 14.4%. The authors conclude that (1) the state of the bone-cement interface as seen on the early postoperative radiograph can predict the longevity of a cemented socket with a high degree of probability, and that (2) cementation of an all-polyethylene socket using modern cement technique is an excellent method of fixation for patients 60 years of age and older with degenerative joint disease.

Proterometra autraini n. sp. (Digenea: Azygiidae) from Michigan's upper peninsula and a key to species of Proterometra.

Life stages of Proterometra autraini n. sp. (Digenea: Azygiidae) from the Au Train River, Au Train, Michigan are described, with emphasis on the cercarial stage. Progenetic cercariae develop in rediae in the snail Elimia livescens and emerge seasonally from mid-summer to mid-autumn. Cercariae emerge naturally between 1000 and 2100 hr EDT. Adults were retrieved from several naturally infected fish species. Specimens of Cottus bairdi (n = 16) displayed the greatest prevalence (75%), mean intensity (7.1), and abundance (5.3) of infection. Experimental infections were successful in C. bairdi, Lepomis macrochirus, and Perca flavescens. The geographical range of this parasite is limited to the portion of the Au Train River between Au Train Lake and Lake Superior.

Lymphocyte subset analysis by Boolean algebra: a phenotypic approach using a cocktail of 5 antibodies and 3 color immunofluorescence.

Commercial reagent kits for the evaluation of leukocyte subsets involve the staining of a panel of up to six tubes using combinations of pre-mixed fluorescein isothiocyanate (FITC) and R-phycoerythrin (PE) conjugated monoclonal antibodies. We describe a rapid method whereby total CD3+ T-cells, CD4+ T-cells (CD3+ CD4+), CD8+ T-cells (CD3+ CD8+), putative gamma delta-receptor-T-cells (CD3+ CD4- CD8-), and T-cells that are CD3+ CD4+ CD8+ as well as B-lymphocytes and NK-cells can be enumerated after staining in a single tube. Whole blood specimens are labelled with a mixture of antibodies: FITC-conjugated antibodies to CD4 and CD19, PE-conjugated antibodies to CD8 and CD16, and either peridinin chlorophyll protein (PerCP) or allophycocyanin (APC) labelling for antibodies to CD3. After recording 20,000 events the data were analysed on the Consort 32 computer system and LYSYS-II (Becton Dickinson, San Jose, CA) and all of the lymphocyte subset values were determined by Boolean algebra using a technique we refer to as Boolean gate analysis (BGA). Our study has shown that BGA is statistically equivalent to SimulSET lymphocyte subset analysis. Furthermore, the procedure reduces the number of tubes required to two with consequential saving in reagents, consumables, and time.