Slow skeletal troponin I gene transfer, expression, and myofilament incorporation enhances adult cardiac myocyte contractile function

The functional significance of the developmental transition from slow skeletal troponin I (ssTnI) to cardiac TnI (cTnI) isoform expression in cardiac myocytes remains unclear. We show here the effects of adenovirus-mediated ssTnI gene transfer on myofilament structure and function in adult cardiac myocytes in primary culture. Gene transfer resulted in the rapid, uniform, and nearly complete replacement of endogenous cTnI with the ssTnI isoform with no detected changes in sarcomeric ultrastructure, or in the isoforms and stoichiometry of other myofilament proteins compared with control myocytes over 7 days in primary culture. In functional studies on permeabilized single cardiac myocytes, the threshold for Ca2+-activated contraction was significantly lowered in adult cardiac myocytes expressing ssTnI relative to control values. The tension-Ca2+ relationship was unchanged from controls in primary cultures of cardiac myocytes treated with adenovirus containing the adult cardiac troponin T (TnT) or cTnI cDNAs. These results indicate that changes in Ca2+ activation of tension in ssTnI-expressing cardiac myocytes were isoform-specific, and not due to nonspecific functional changes resulting from overexpression of a myofilament protein. Further, Ca2+-activated tension development was enhanced in cardiac myocytes expressing ssTnI compared with control values under conditions mimicking the acidosis found during myocardial ischemia. These results show that ssTnI enhances contractile sensitivity to Ca2+ activation under physiological and acidic pH conditions in adult rat cardiac myocytes, and demonstrate the utility of adenovirus vectors for rapid and efficient genetic modification of the cardiac myofilament for structure/function studies in cardiac myocytes.     

Prosthetic valve endocarditis: superiority of surgical valve replacement versus medical therapy only

The objective of our study was to assess the long-term outcome of patients with prosthetic valve endocarditis. We used a multicenter, prospective, observational study design. Six university teaching hospitals with high volume cardiothoracic surgery participated. Seventy-four patients with prosthetic valve endocarditis as defined by explicit, objective criteria were selected for participation. All patients were followed up prospectively for 1 year. Thirty-one percent and 69% had development of endocarditis within 60 days of valve insertion ("early") and after 60 days ("late"), respectively. The most common causes were Staphylococcus epidermidis (40%), Staphylococcus aureus (20%), streptococcal species (18%), and aerobic gram-negative bacilli (11%). Physical signs of endocarditis (new or changing murmur, stigmata, emboli) were seen in 58%. At 6 months and 12 months, mortality was 46% and 47%, respectively. Surgical replacement of the infected valve led to significantly lower mortality (23%) as compared with medical therapy alone (56%), as assessed by both univariate and multivariate analyses (p < 0.05). Improved outcome was seen for the surgical group even when controlling for severity of illness at time of diagnosis. From these findings we conclude that accurate assessment of outcome in prosthetic valve endocarditis requires long-term follow-up of at least 6 months following diagnosis. Surgical therapy warrants greater scrutiny; evaluation in controlled clinical trials is appropriate.     

Biology of chronic radiation effect on tissues and wound healing

Radiation therapy exerts both acute and chronic effects on normal tissue included within treatment fields. The physics of radiation therapy and treatment techniques to minimize deleterious effects of radiation are presented. Management of radiation-damaged skin is discussed. Radiation effect on tissue, wound healing, and tumorigenesis also are reviewed.     

The B-cell transmembrane protein CD72 binds to and is an in vivo substrate of the protein tyrosine phosphatase SHP-1

BACKGROUND: Signals from the B-cell antigen receptor (BCR) help to determine B-cell fate, directing either proliferation, differentiation, or growth arrest/apoptosis. The protein tyrosine phosphatase SHP-1 is known to regulate the strength of BCR signaling. Although the B-cell co-receptor CD22 binds SHP-1, B cells in CD22-deficient mice are much less severely affected than those in SHP-1-deficient mice, suggesting that SHP-1 may also regulate B-cell signaling by affecting other signaling molecules. Moreover, direct substrates of SHP-1 have not been identified in any B-cell signaling pathway. RESULTS: We identified the B-cell transmembrane protein CD72 as a new SHP-1 binding protein and as an in vivo substrate of SHP-1 in B cells. We also defined the binding sites for SHP-1 and the adaptor protein Grb2 on CD72. Tyrosine phosphorylation of CD72 correlated strongly with BCR-induced growth arrest/apoptosis in B-cell lines and in primary B cells. Preligation of CD72 attenuated BCR-induced growth arrest/death signals in immature and mature B cells or B-cell lines, whereas preligation of CD22 enhanced BCR-induced growth arrest/apoptosis. CONCLUSIONS: We have identified CD72 as the first clear in vivo substrate of SHP-1 in B cells. Our results suggest that tyrosine-phosphorylated CD72 may transmit signals for BCR-induced apoptosis. By dephosphorylation CD72. SHP-1 may have a positive role in B-cell signaling. These results have potentially important implications for the involvement of CD72 and SHP-1 in B-cell development and autoimmunity.     

Identification of a RAPD marker linked to progressive rod-cone degeneration in dogs

1. Tammar Wallaby embryonic blood has been shown to have three alpha-like and two beta-like globin chains in its four haemoglobin components and partial sequences of several chains have been determined. 2. The major embryonic beta-like chain (epsilon) is similar to other mammalian embryonic beta-like chains on the basis of sequencing its first 60 amino acids. 3. There is another embryonic beta-like chain present in one haemoglobin component. It was designated omega and, in its first 54 amino acids, it has features that are more like avian globins than mammalian globins. 4. The one alpha-like embryonic globin sequenced has mammalian rather than avian characteristics. 5. A provisional phylogenetic tree of beta-like globins has been determined. The Tammar epsilon-globin forms a monophyletic group with marsupial and other mammalian embryonic globins; the omega-globin forms a monophyletic group with bird adult and embryonic globins.     

Glycopeptide antibiotic resistance genes in glycopeptide-producing organisms

The mechanism of high-level resistance to vancomycin in enterococci consists of the synthesis of peptidoglycan terminating in D-alanyl-D-lactate instead of the usual D-alanyl-D-alanine. This alternate cell wall biosynthesis pathway is ensured by the collective actions of three enzymes: VanH, VanA, and VanX. The origin of this resistance mechanism is unknown. We have cloned three genes encoding homologs of VanH, VanA, and VanX from two organisms which produce glycopeptide antibiotics: the A47934 producer Streptomyces toyocaensis NRRL 15009 and the vancomycin producer Amycolatopsis orientalis C329.2. The predicted amino acid sequences are highly similar to those found in VRE: 54 to 61% identity for VanH, 59 to 63% identity for VanA, and 61 to 64% identity for VanX. Furthermore, the orientations of the genes, vanH, vanA, and vanX, are identical to the orientations found in vancomycin-resistant enterococci. Southern analysis of total DNA from other glycopeptide-producing organisms, A. orientalis 18098 (chloro-eremomycin producer), A. orientalis subsp. lurida (ristocetin producer), and Amycolatopsis coloradensis subsp. labeda (teicoplanin and avoparcin producer), with a probe derived from the vanH, vanA, and vanX cluster from A. orientalis C329.2 revealed cross-hybridizing DNA in all strains. In addition, the vanH, vanA, vanX cluster was amplified from all glycopeptide-producing organisms by PCR with degenerate primers complementary to conserved regions in VanH and VanX. Thus, this gene sequence is common to all glycopeptide producers tested. These results suggest that glycopeptide-producing organisms may have been the source of resistance genes in vancomycin-resistant enterococci.     

Genetic diversity among Mycobacterium avium complex strains recovered from children with and without human immunodeficiency virus infection

The genetic diversity and molecular epidemiology of Mycobacterium avium complex (MAC) infections in children with and without human immunodeficiency virus (HIV) infection were evaluated. Isolates recovered from 136 children were subtyped by sequence analysis of a 360-bp region of the gene (hsp65) encoding a 65-kDa heat-shock protein. Twenty-one distinct hsp65 alleles were identified. On the basis of hsp65 genotype, 6 isolates were not MAC organisms. Of the remaining 130 samples, 61% were M. avium, 37% were Mycobacterium intracellulare, and 2% were species nonspecific MAC. Eighty-eight percent of the isolates obtained from HIV-infected children were M. avium. In contrast, only 38% of the isolates obtained from children without HIV infection were M. avium (chi2 test, P < .001). M. avium isolates were further subtyped by Southern blot analysis with insertion element IS1245. Taken together, no evidence for a single clonal M. avium strain causing infection was detected.     

Differing haemodynamic and catecholamine responses to exercise in three groups with peripheralautonomic dysfunction: insulin-dependent diabetes mellitus, familial amyloid polyneuropathy and pure autonomic failure

BACKGROUND: The nosological status of postpartum psychoses has remained controversial because of their often 'atypical' symptomatology. A polydiagnostic approach may further clarify this issue. METHODS: In a retrospective study, we applied the ICD-10 and Leonhard's classification to 39 patients with severe postpartum psychiatric disorders. The patients were personally reexamined on average 12.5 years (6-26 years) after the onset of the illness. RESULTS: An acute onset and a polymorphous psychotic symptomatology with rapid changes characterized the majority of our cases. Unipolar depressive disorders (28%) and acute polymorphous psychotic disorders (21%) represented the largest proportions within the ICD-10-classification. Applying Leonhard's classification, over half the patients (54%) suffered from a cycloid psychosis. Among cycloid psychoses, motility psychoses clearly predominated. Schizophrenias occurred rarely (10%) according to both classifications. LIMITATIONS: Due to the unknown prevalence of the various diagnoses among women of child-bearing age, it is impossible to statistically infer a specific association between childbirth and a distinct diagnosis from our data. CONCLUSIONS: Our findings suggest that cycloid psychoses, in particular motility psychoses, account for the majority of postpartum psychoses, and do not support the hypothesis of a nosological independence of postpartum psychoses.     

The humoral response to xenografts is controlled by a restricted repertoire of immunoglobulin VH genes

BACKGROUND: The early phases of the host immune response to xenografts are dominated by anti-donor antibodies. The immunological pathways responsible for mediating the host humoral responses to xenografts are largely unknown, and this report addresses the nature of the immunoglobulin genes controlling the host antibody response to xenografts. METHODS: cDNA libraries established from rat anti-hamster monoclonal antibodies and splenic lymphocytes from LEW rats rejecting hamster heart xenografts were used to clone, sequence, and identify the immunoglobulin genes responsible for encoding rat xenoantibodies to hamster heart grafts. Libraries for germline variable region heavy chain (VH) genes encoding the anti-hamster xenograft antibodies were established by genomic DNA cloning and analyzed by nucleotide sequencing. The frequency of Ig VH gene usage for controlling the antibody responses to hamster xenografts was examined by colony-filter dot hybridization. The nucleic acid structure of these genes was then compared to their genomic progenitors to identify the number and structural diversity expressed by the Ig VH genes used to mediate the response. RESULTS: Rat monoclonal antibodies selected for their ability to precipitate the rejection of hamster xenografts exclusively use a closely related group of VH genes. The VH genes used by these antibodies are restricted to a single family of germline genes (VHHAR) for which 15 family members have been identified. The frequency of VHHAR gene usage in splenic IgM-producing B cells from LEW rats rapidly expands from 0.8% in naive animals to 13% in recipients 4 days after xenotransplantation. cDNA libraries expressing VHHAR genes were established from splenic lymphocytes derived from naive or xenograft recipients at 4 and 21 days after transplantation. Examination of 20 cDNA clones revealed that the majority (75%) of these clones express VHHAR genes displaying limited somatic mutation. CONCLUSIONS: The use of a closely related group of Ig VH genes in a germline configuration to control the early humoral response to xenografts suggests that this response may represent the utilization of a primitive, T cell-independent pathway of antibody production by the graft recipients.