Apoptosis and apoptosis-associated perturbations of peripheral blood lymphocytes during HIV infection: comparison between AIDS patients and asymptomatic long-term non-progressors

This study was designed to compare the degree of lymphocyte apoptosis and Fas-Fas ligand (FasL) expression in AIDS patients and long-term non-progressors (LTNPs) and correlate these parameters with apoptosis-associated perturbations in lymphocyte function. LTNPs had a lower frequency of apoptotic CD4+ and CD8+ T cells compared with subjects with AIDS. This correlated with a lower frequency of cells expressing Fas and FasL. The frequency of selected lymphocyte populations exhibiting a disrupted mitochondrial transmembrane potential (DeltaPsim) and increased superoxide generation was lower in LTNPs than in patients with AIDS; these abnormalities were associated with lower levels of caspase-1 activation in LTNPs. The results indicate a significantly reduced level of apoptosis and apoptosis-associated parameters in LTNPs than in patients developing AIDS. Based on these findings, a crucial role for mitochondria can be predicted in the process of lymphocyte apoptosis during the evolution of AIDS.     

Antibodies to nucleic acids in myasthenia gravis.

An increased frequency of antibodies to native DNA, thymocytes, and striated muscle was found in patients with myasthenia gravis (MG). The prevalence of such antibodies lends considerable support to the concept of MG as an autoimmune disorder and militates in favor of major abnormalities in the thymic dependent immune system. There was no correlation between serum-blocking activity to acetylcholine receptor protein and antibodies to thymocytes.     

Alkaline phosphatase isoenzymes and osteocalcin in serum of normal subjects

Clinical laboratory tests are increasingly being used to evaluate individuals for osteoporosis and other metabolic bone diseases. Serum bone alkaline phosphatase (AP) [EC 3.1.3.1, orthophosphoric-monoester phosphohydrolase (alkaline optimum)] and osteocalcin are used to assess osteoblastic activity. Although methods for assessing relative amounts of AP isoenzymes continuously appear in the literature, no single method is satisfactory for quantification. Polyacrylamide gel electrophoresis with densitometric scanning combined with two-point heat inactivation was used to obtain quantitative values for AP isoenzymes. Serum bone AP concentrations correlated positively and significantly with serum osteocalcin concentrations obtained by radioimmunoassay for women. Men had significantly higher total alkaline phosphatase and bone AP than women, whereas liver AP concentrations did not differ between the two groups. Bone AP correlated negatively and significantly with age in men, but not women. Osteocalcin concentrations tended to be higher in men, but not significantly.     

Electrical stimulation in the treatment of osteonecrosis of the femoral head--a 1-year follow-up

To date, there is no completely satisfactory method for the treatment of osteonecrosis of the femoral head. "Conservative" management leads to a high failure rate, and surgical results have been inconsistent and disappointing. The study described in this article seeks to evaluate the role of electrical stimulation in conjunction with decompression and bone grafting. A total of 82 hips have been included to date, of which 42 with a minimum follow-up of 1 year have been evaluated. Pain relief and improved function have been noted in the majority of hips operated on, both with and without electrical stimulation. Careful radiologic assessment using a comprehensive new method of evaluation has shown some degree of progression in the majority of cases, however. No effects of electrical stimulation per se have been demonstrated to date. It must be emphasized that this is a preliminary report with a minimum follow-up of 1 year. Final conclusions await the completion of the study, when all patients have been followed for a minimum of 3 years.     

Isolation of the non-glycosylated proteins of desmosomes and immunolocalization of a third plaque protein: desmoplakin III.

The cytoplasmic plaque of the spot desmosome or macula adhaerens mediates the attachment of bundles of intermediate filaments to the plasma membrane. We have isolated from a bovine epidermal desmosome preparation a fraction that is highly enriched in the non-glycosylated desmosomal proteins. Plastic-embedded and thin-sectioned high-speed pellets of this fraction reveal closely packed filaments that resemble plaque regions of the low pH whole desmosome preparation from which they are derived. NaDodSO4/polyacrylamide gel electrophoresis reveals four major, non-glycosylated proteins of 240, 210, 81, and 77 kDa. In agreement with a previous study, we find the 240- and 210-kDa proteins (desmoplakins I and II) to be closely related, whereas the 81- and 77-kDa proteins are unique. This is shown both immunologically and by one-dimensional proteolytic peptide mapping. Monospecific, polyclonal rabbit antibodies were prepared against the 81-kDa protein and used, in conjunction with protein A-complexed colloidal gold particles (PAG), to immunolocalize this antigen on ultrathin sections of bovine muzzle epidermis. On antibody-labeled sections, PAG particles were associated principally with the desmosomal cytoplasmic plaque. Sections exposed to preimmune serum showed little or no labeling. We conclude that the 81-kDa protein, like the 240/210-kDa protein family, is one of the major components of the desmosomal plaque. We designate it as "desmoplakin III." The location of the 77-kDa protein remains to be definitively established.     

Epidermal cell-shape regulation and subpopulation kinetics during butyrate-induced terminal maturation of normal and SV40-transformed human keratinocytes: epithelial models of differentiation therapy

Recent data indicate that malignant human epidermal cells may be appropriate targets for sodium butyrate (NaB)-mediated differentiation therapy. The response of pre- and post-crisis populations of SV40-transformed human keratinocytes (SVKs) to this differentiation-inducing agent was assessed, therefore, within the framework of NaB-directed normal human keratinocyte (NHK) maturation. NaB augmented cornified envelope (CE) production in NHK and pre-crisis SVK cultures; the time-course and efficiency of induced maturation were similar in the 2 cell systems. In NHKs, the percentage of amplifying ("B" substate) cells decreased with time in NaB correlating with increases in both "C" stage keratinocytes and CEs. The latter formed over one or 2 layers of nucleated basal-like cells. Inductions were accompanied by immediate cell cycle blocks (in both the G1 and G2/M phases), reorganization within the actin cytoskeleton, and transient early increases in cellular actin content. Increased NHK and pre-crisis SVK cytoskeletal-associated actin reached a maximum approximately 48 hr after NaB addition and preceded development of CEs. The CE precursors, thus, probably reside in the "B" substate. Post-crisis SVKs, in contrast, were refractive to NaB-induced terminal maturation or cell-cycle perturbation, failed to initiate actin filament rearrangements, and retained a basal cell-like phenotype. Stable transformation of human SVKs in post-crisis phase, therefore, appears to be associated with loss of maturation "competence" within the "B" keratinocyte subpopulation.     

Expression of pyruvate kinase M2 in preneoplastic hepatic foci of N-nitrosomorpholine-treated rats

 The expression of the pyruvate kinase (PK) isoenzymes L and M₂ was analysed in the livers of rats treated with the hepatocarcinogenic agent N-nitrosomorpholine (NNM) in the drinking water. In control animals L-PK expression was restricted to liver parenchymal cells, whereas M₂-PK was detected in bile duct epithelial, blood vessel wall, endothelial and Kupffer cells. In rats treated with NNM proliferating oval cells were consistently L-PK negative and M₂-PK positive, while the ductal cells of cholangiofibroses were clearly L-PK positive and coexpressed M₂-PK. However, no morphological differentiation of ductal cells into hepatocyte-like cells was observed. In the clear and acidophilic cell foci storing glycogen in excess strong staining for L-PK was observed. In glycogen-poor foci induced by NNM a shift from L-PK to M₂-PK expression takes place. Received: 24 March 1998 / Accepted: 13 November 1998     

Recurrent respiratory papillomatosis: altered CD8(+) T-cell subsets and T(H)1/T(H)2 cytokine imbalance

Human papillomaviruses (HPVs) cause benign papillomas and squamous cell carcinomas in the genital and respiratory tracts. Recurrent respiratory papillomas (RRP) generate a high level of morbidity and significant mortality because of their location, resistance to treatment, and relentless recurrence that can vary in frequency in a given patient and between patients. We have found that T-cells from these patients, when exposed to or isolated from autologous papilloma tissue, have an elevated percentage of CD8(+), CD28(-) T-cells, and that T-cells from many of these patients express an increase in T(H)2-like cytokine mRNA in response to autologous papilloma tissue. Furthermore, both of these immunologic findings correlate with disease severity. These observations suggest that patients with RRP, and possibly others with refractory HPV-induced lesions, are unable to manage their disease with an appropriate and effective HPV-specific, T-cell response. This immune imbalance may be responsible for the development and severity of HPV-induced respiratory papillomatosis.     

Gene therapy using viral vectors for acute neurologic insults

Enormous knowledge has emerged concerning the cellular and molecular events underlying necrotic neuron death after seizure, hypoxia-ischemia, or hypoglycemia. This has allowed the design of rational therapies to protect neurons at such times. One of the most exciting arenas of such interventions is the use of viral vectors to deliver neuroprotective genes. This review considers the progress in this nascent discipline. Neuroprotection has been demonstrated against a variety of in vitro and in vivo rodent models of necrotic insults with vectors overexpressing genes that target various facets of injury. These have included the energetic components, calcium excess, accumulation of reactive oxygen species, protein malfolding, inflammation, and triggering of apoptosis (i.e., programmed cell death) in a subset of cells. A number of caveats, subtleties, and pressing questions concerning this literature then are considered. These include whether these gene therapy interventions actually prevent, rather than merely delay, neuron death; the extent to which the effects of such vectors on neuronal cell biology is actually understood; the potential adverse effects of the use of such vectors; and whether sparing a neuron from death with one of these interventions spares function as well. Finally, we consider the likelihood of such gene therapy becoming relevant to clinical neurology in the near future.