Identification and characterization of alternatively spliced murine Rgs11 isoforms: genomic structure and gene analysis.

The RGS proteins comprise a large family of proteins which were recently identified as negative Regulators of G-protein Signaling. They have been shown to act as GTPase Activating Proteins (GAPs) towards the G(alpha) subunits of heterotrimeric G-proteins. In addition to this GAP activity, which has been shown to occur through the RGS domain, RGS proteins are likely to possess other functions due to the existence of other domains in these molecules (De Vries and Farquhar, 1999; Hepler, 1999). Here, we report the molecular characterization of the murine Rgs11 gene. The gene encodes a protein with high homology to human RGS11 (79.9%), containing conserved DEP (Dishevelled/EGL-10/Pleckstrin) and GGL (G protein gamma-like) domains. The gene is comprised of at least 13 exons, spanning 8-9 kb. Spliced transcript variants were identified which are co-expressed with 5A3, a transcript that contains the largest ORF. Expression of mouse Rgs11 was found to be restricted to specific tissues with a unique pattern of expression observed in brain.

Blood biochemical profile of the South American rattlesnake (Crotalus durissus terrificus) in captivity

Blood samples were collected from 180 healthy specimens of the South American rattesnake, Crotalus durissus terrificus, in captivity. All animals were in good clinical condition. Normal biochemical reference values were established for the following: total proteins, albumin, globulins, uric acid, creatinine, urea, glucose, triglycerides, cholesterol, total lipids, calcium, phosphorus, potassium, sodium, chloride, magnesium, GOT (AST), GPT (ALT), and alkaline phosphatase (ALP). Samples were obtained by venipuncture of the ventral tail vein. Values were compared with published data for Boidae, Elapidae, and Viperidae.

Methylation-dependent silencing of the testis-specific Pdha-2 basal promoter occurs through selective targeting of an activating transcription factor/cAMP-responsive element-binding site.

In this study, we demonstrate that methylation-dependent repression of the Pdha-2 core promoter is mediated regionally through a consensus activating transcription factor/cAMP-responsive element-binding site located between nucleotides -54 and -62 upstream of the major transcriptional start site. Targeting of the CpG dinucleotide within this cis-element significantly disrupts the ability of this basal promoter to activate gene expression in vitro and completely abolishes promoter activity in vivo. DNase I footprinting experiments indicated that availability of the nuclear factor(s) binding this element is limiting in sexually immature mouse testis, and as such, these factors may play an important role in the coordinate activation of early spermatogenic gene expression. Interestingly, CpG dinucleotides associated with the hypersensitive region flanking the activating transcription factor/cAMP-responsive element-binding site appear to confer some conformational structure on the promoter since mutations at these specific CpG dinucleotides result in elevated basal levels of transcription. This raises the possibility of a potential bifunctional role for CpG dinucleotides in either methylation-dependent or -independent processes. Our data support the notion that hypomethylation and transcription factor recruitment are necessary events that precede gene activation at the early stages of spermatogenesis.

Review: Pdha-2, past and present.

Pyruvate dehydrogenase (PDH) is a multiunit enzymatic complex essential for the process of generating cellular energy. One of the most important of its subunits is the E1 alpha subunit. Perturbations in the expression of this subunit lead to reduced or lost function of the PDH complex as a whole, resulting in a loss of ATP production. The consequence of such perturbations can lead to neurological abnormalities, lactic acidosis, and in males, death. Pdha-2 codes for the mouse testis isoform of the E1 alpha subunit and maps to chromosome 19 (chromosome 4 in humans). This is a fortuitous evolutionary development because the somatic isoform of the E1 alpha subunit is linked to the X-chromosome, which is not only inactivated early in spermatogenesis but is represented in only half of the haploid spermatid population. Consequently, activation of the testis-specific E1 alpha subunit is essential for the progression of spermatogenesis. Despite its importance, the molecular mechanisms governing the tight tissue- and temporal-specific regulation of Pdha-2 have, until recently, remained poorly understood. In this review, we describe our current understanding of the transcriptional regulation of Pdha-2 and propose potential mechanisms that may play a role in this process.

The Editor's Role in the Science and Art of Physical Therapy.

James A. Gould, MS, PT, delivered the Paris Distinguished Service Award Lecture at the Combined Sections meeting in San Francisco in February. Gould is the second recipient of the Orthopaedic Section's Paris Award, which was named for its first recipient and the founder of the Orthopaedic Section, Stanley Paris, PhD, PT. In 1968, Gould graduated from Central Michigan University with a BS degree in biology. He earned a BS degree in physical therapy from the University of Kentucky in 1971 and a master's degree in education from the University of Kentucky in 1974. Gould was then appointed a faculty member of the University of Wisconsin-La Crosse, where he is now an associate professor. In 1979, Gould founded The Journal of Orthopaedic and Sports Physical Therapy and served as editor until 1990. He is currently treasurer of the Private Practice Section, editor of PT Today, and co-owner of OPTions, a clinical practice in La Crosse, WI. J Orthop Sports Phys Ther 1992;15(6):323-325.

Interleukin-2 and high-dose cisplatin in patients with metastatic melanoma: a pilot study.

In this pilot study of metastatic melanoma, interleukin-2 (IL-2) and cisplatin (CDDP) chemotherapy were combined using an alternating schedule designed to explore potential synergism between these modalities. Bolus IL-2 was given at a dose of 600,000 IU/kg intravenously (IV) every 8 hours, days 1 to 5 and 15 to 19, followed by high-dose CDDP administered by two different regimens: (A) 135 to 150 mg/m2 IV bolus over 30 minutes with the chemoprotectant WR-2721 910 mg/m2 or (B) 50 mg/m2 IV over 2 hours every day for 3 days. The trial design allowed an assessment of response to each phase of therapy. Among 27 assessable patients, there were 10 (37%) overall responses, including three (11%) complete responses (CRs) with durations of 9, 16, and 30+ months. Tumor regression was noted in seven patients (partial response [PR], four; minor response [MR], three; response rate [RR], four of 27 [15%]) after IL-2 administration and in 14 patients (PR, 12; MR, two; RR, 12 of 27 [44%]) after CDDP treatment, demonstrating noncrossresistance between the components of the regimen. Major PRs (greater than 90% reduction of tumor burden) or CRs were only seen in patients responding to IL-2. Toxicity during IL-2 therapy was typical for high-dose IL-2 protocols and was reversible. Among the first 20 patients treated with CDDP regimen A, there were eight episodes of grade IV nephrotoxicity (creatinine level greater than 5.0 mg/dL), including three of six patients treated with an initial CDDP dose of 135 mg/m2. This side effect was more frequent among patients with liver metastasis (P less than .05, Fisher's exact test). No significant nephrotoxicity was noted in seven patients treated on regimen B. Although ototoxicity was frequent, minimal bone marrow and neurologic toxicity was noted. There were no treatment-related deaths. This combination showed at least additive activity against melanoma, and the more protracted CDDP schedule was well tolerated. This regimen may serve as a model for future combined immunotherapy and chemotherapy trials in metastatic melanoma.

Procurement and transplantation of colonized cadaver skin.

Cadaver skin is an important adjunct to burn care. This study was designed to measure the percentage of its contamination prior to grafting and to determine the clinical safety of its use. Cadaveric homografts from 19 donors were harvested, frozen, thawed, and used as a biological dressing. Cultures were obtained at the time of harvest, after incubation in antibiotics, and at actual transplantation. The homografts were 43 per cent, 68 per cent, and 50 per cent contaminant-free, respectively. Two of 14 patients lost their homografts due to infection. However, no patient developed an infection or lost the homograft due to an organism identified during the pretransplantation processing. Therefore, contaminant-free and slightly contaminated cadaver skin can be used safely as homografts.

Nosocomial sepsis associated with interleukin-2.

STUDY OBJECTIVE: To determine the incidence, clinical magnitude, and risk factors for nosocomial bacteremia in patients given interleukin-2 with or without (+/-) lymphokine activated killer (LAK) cells for cancer immunotherapy. DESIGN: Cohort study. SETTING: Clinical study unit of tertiary medical center. PATIENTS: All patients entering the interleukin-2 +/- LAK cancer immunotherapy protocol during a 28-month period. Control groups were patients in a surgical intensive care unit, patients receiving total parenteral nutrition, and patients with solid tumors. MEASUREMENTS AND MAIN RESULTS: Twenty of 107 (19%) interleukin-2-treated patients developed sepsis; in 12 of these patients, sepsis was intravenous catheter-associated. The bacteremia rate among patients receiving total parenteral nutrition, in the surgical intensive care unit, or having solid tumors was 2.8%, 4.1%, and 1.9%, respectively. Staphylococcus aureus was the pathogen in 13 courses; Staphylococcus epidermidis, in 5; and Escherichia coli, in 2. Two patients died; three developed suppurative thrombophlebitis; one developed septic arthritis; one, septic arterial aneurysm; and one, peritonitis with probable meningitis. Colonization with S. aureus increased the risk of S. aureus bacteremia 6.3-fold (95% CI, 2.8 to 14.5; P less than 0.001); skin desquamation at the catheter site increased the relative risk 2.0-fold (95% CI, 1.3 to 3.1; P = 0.031). Both colonization with S. aureus and skin desquamation increased the relative risk of S. aureus bacteremia 14.5-fold (95% CI, 4.1 to 50.9; P less than 0.0001). CONCLUSIONS: Staphylococcal bacteremia is more frequent in patients receiving interleukin-2 therapy and is associated with substantial morbidity and toxic skin reactions.