Immunohistological characterization of spinal TB granulomas from HIV-negative and -positive patients.

Tuberculosis (TB) is mainly a disease of the lungs, but Mycobacterium tuberculosis (Mtb) can establish infection in virtually any organ in the body. Rising rates of extrapulmonary (EP) TB have been largely associated with the HIV epidemic, as patients co-infected with HIV show a four-fold higher risk of EPTB. Spinal TB (Pott's Disease), one of the most debilitating extrapulmonary forms of disease, is difficult to diagnose and can cause deformity and/or neurological deficits. This study examined the histopathology and distribution of immune cells within spinal TB lesions and the impact of HIV on pathogenesis. The overall structure of the spinal granulomas resembled that seen in lung lesions from patients with pulmonary TB. Evidence of efficient macrophage activation and differentiation were detectable within organized structures in the spinal tissue, irrespective of HIV status. Interestingly, the granulomatous architecture and macroscopic features were similar in all samples examined, despite a reversal in the ratio of infiltrating CD4 to CD8 T cells in the lesions from HIV-infected patients. This study provides a foundation to understand the mechanism of tissue destruction and disease progression in Spinal TB, enabling the future development of novel therapeutic strategies and diagnostic approaches for this devastating disease.

Mutations in the epsilon sequences of human hepatitis B virus affect both RNA encapsidation and reverse transcription.

Hepadnaviruses replicate by reverse transcription of an RNA intermediate within subviral core particles in the cytoplasm of infected hepatocytes. Recognition of the epsilon encapsidation signal located on the 5' end of the pregenomic RNA by the viral polymerase occurs early in core particle assembly. The epsilon sequences contain a set of nested inverted repeats which form a stable stem-loop structure shown to play a role in RNA packaging and recently implicated as the site of initiation of minus-strand DNA synthesis. We have introduced a variety of site-directed mutations into the epsilon sequences of human hepatitis B virus to study their effects on viral replication in transfected HuH7 cells. We have identified two classes of mutations: those which adversely affect packaging and those which package RNA but adversely affect DNA synthesis. Analysis of these mutants has allowed us to identify separate features of the epsilon cis-acting signal which function in the processes of RNA packaging and reverse transcription.

Genes encoding alpha and beta subunits of Na,K-ATPase are located on three different chromosomes in the mouse.

We have made use of a panel of mouse-hamster somatic cell hybrids and restriction fragment length polymorphisms between two mouse species (Mus musculus and Mus spretus) to determine the chromosomal localization of genes encoding the alpha and beta subunits of the Na,K-ATPase (Na+,K+-activated ATP phosphohydrolase, EC 3.6.1.3). DNA probes for three distinct isoforms of the Na,K-ATPase alpha subunit mapped to three different mouse chromosomes: the alpha 1 gene (Atpa-1) cosegregated with the Egf gene on chromosome 3; alpha 2 (Atpa-2) with the cytochrome P-450PB gene family/coumarin hydroxylase locus on chromosome 7; alpha 3 (Atpa-3) with the alpha-spectrin gene on chromosome 1. The Na,K-ATPase beta-subunit gene (Atpb) mapped to the same region of chromosome 1, but it was not tightly linked to the Atpa-3 gene. These results indicate that three isoforms of the Na,K-ATPase alpha subunit are encoded by three distinct genes. The dispersion of Na,K-ATPase genes suggests that their expression is not likely to be controlled by a common cis-acting regulatory element.

Chromosome-mediated gene transfer of multidrug resistance.

Multidrug resistance can be transferred from drug-resistant LZ Chinese hamster cells to drug-susceptible mouse LTA cells by chromosome-mediated gene transfer. Analysis of genomic DNA demonstrated the transfer of multiple copies of a DNA domain which is amplified in the donor multidrug-resistant cells. The transfer of 10 to 15 copies of the Chinese hamster gene was sufficient to produce a multidrug-resistant phenotype. Chromosome transferents exhibited overexpression of an mRNA of approximately 5 kilobases which has previously been demonstrated to be encoded by the amplified DNA domain of the donor LZ cells. Phenotypic analysis of individual clones selected in adriamycin showed the resistance to be pleiotropic. All clones tested demonstrated similar levels of cross-resistance to the drugs daunorubicin and colchicine. These results indicate that the DNA sequences transferred confer the complete multidrug-resistant phenotype on recipient cells and suggest that multidrug resistance is due to overexpression of the protein encoded by the 5-kilobase mRNA.