Restoration of cytomegalovirus-specific CD4+ T-lymphocyte responses after ganciclovir and highly active antiretroviral therapy in individuals infected with HIV-1.

Recent studies of subjects infected with human immunodeficiency virus (HIV-1) have produced conflicting results about the extent of reconstitution possible in the CD4+ lymphocyte repertoire after highly active antiretroviral therapy (HAART). The effect of HAART on the incidence of opportunistic infections will probably depend on reconstitution of antigen-specific CD4+ lymphocyte responses to important pathogens, including cytomegalovirus (CMV), the leading cause of blindness in AIDS. Several studies have demonstrated an important role for CD4+ lymphocytes in controlling CMV replication in vitro and in clinical studies. It is now possible to quantitate antigen-specific CD4+ lymphocyte responses by flow cytometry. Using this method, we studied CMV-specific CD4+ lymphocyte responses in individuals infected with HIV-1 with and without a history of active CMV-associated end organ disease (EOD), and in those with quiescent CMV EOD after ganciclovir therapy and HAART. The presence of active CMV-associated EOD strongly correlated with loss of CMV-specific lymphocyte responses (P = 0.0004). In contrast, patients with no history of CMV-associated EOD and most patients with quiescent EOD after HAART demonstrated strong CMV-specific CD4+ lymphocyte responses. These data indicate that the loss of CMV-specific CD4+ lymphocyte responses in individuals infected with HIV-1 who have active CMV EOD may be restored after ganciclovir therapy and HAART, which provides evidence for functional immune reconstitution to an important pathogen.

Induction of AIDS-like disease in macaque monkeys with T-cell tropic retrovirus STLV-III.

The T-cell tropic retrovirus of macaque monkeys STLV-III has morphologic, growth, and antigenic properties indicating that it is related to HTLV-III/LAV, the etiologic agent of the acquired immune deficiency syndrome (AIDS) in humans. Four of six rhesus monkeys died within 160 days of STLV-III inoculation with a wasting syndrome, opportunistic infections, a primary retroviral encephalitis, and immunologic abnormalities including a decrease in T4+ peripheral blood lymphocytes. These data show that an immunodeficiency syndrome can be produced experimentally in a nonhuman primate by an agent from the HTLV-III/LAV group of retroviruses. The STLV-III-macaque system will thus provide a useful model for the study of antiviral agents and vaccine development for human AIDS.

High prevalence of thymic tissue in adults with human immunodeficiency virus-1 infection.

The thymus in adults infected with the HIV-1 is generally thought to be inactive, both because of age-related involution and viral destruction. We have revisited the question of thymic function in adults, using chest-computed tomography (CT) to measure thymic tissue in HIV-1-seropositive (n = 99) or HIV-1-seronegative (n = 32) subjects, and correlating these results with the level of circulating CD4(+) and CD8(+) T cells that are phenotypically described as naive thymic emigrants. Abundant thymic tissue was detectable in many (47/99) HIV-1-seropositive adults, aged 20-59. Independent of age, radiographic demonstration of thymic tissue was significantly associated with both a higher CD4(+) T cell count (P = 0.02) and a higher percentage and absolute number of circulating naive (CD45RA+CD62L+) CD4(+) T cells (P < 0.04). The prevalence of an abundant thymus was especially high in younger HIV-1-seropositive adults ( 40 yr) regardless of CD4 count (P = 0.03). These studies suggest that the thymus is functional in some but not all adults with HIV-1 disease.

Standardized neurologic evaluations of 128 patients with Wegener granulomatosis.

OBJECTIVE: To assess the frequency and type of neurologic involvement in a cohort of patients with generalized Wegener granulomatosis (WG). PATIENTS AND METHODS: In a prospective analysis the clinical, electrophysiologic, radiological, and serologic data of 128 patients have been studied over a median observation period of 19 months (range, 1-60 months). RESULTS: Sixty-four patients (50%) revealed central or peripheral nervous system involvement. Peripheral neuropathy (PN) affected 56 patients, in 9 cases the central nervous system was involved, and in 6 cases the cranial nerves were involved. Thirty-one patients showed a distal symmetrical polyneuropathy, 25 a mononeuritis multiplex. Within the first 2 years of the disease course 47 of the 56 patients had developed their PN, sometimes as the initial symptom of WG. Patients with PN were significantly more often male (34 of 65 patients) than female (22 of 63 patients, P =.04), were significantly older at the onset of WG (median age, 53 vs 44 years; P =.001), had a significantly larger disease extent (P =.001), and had higher classic antineutrophil cytoplasmic antibody titers (P =.002) than neurologically unaffected patients. Response to immunosuppression was moderate concerning peripheral nervous system manifestations. CONCLUSIONS: Peripheral neuropathy is frequent in generalized WG, occurring early in the disease course. As PN can be the first and sole symptom of a beginning systemic vasculitis, it is important that in cases of PN of an unclear origin, interdisciplinary investigations are initiated to detect, treat, and closely follow-up a possible underlying WG, especially as these patients seem to have a more severe disease course.

Evaluation of sympathetic blockade after intrathecal and epidural lidocaine in rats by laser Doppler perfusion imaging.

The widespread use of neuraxial anaesthesia increases the need for animal models to evaluate therapeutic prospects, mechanisms and risks of this technique. As a methodological prerequisite, we characterised the sympathetic blockade after different modes of neuraxial anaesthesia with regard to segments supplying the splanchnic region. Under haemodynamic monitoring, lidocaine 2% or saline were infused via intrathecal (10 microl), lumbar epidural (10 and 30 microl) or thoracic epidural (10 and 30 microl) catheters. Segmental spread of neuraxially infused local anaesthetic was assessed using methylene blue. Mean arterial blood pressure decreased more severely after neuraxial lidocaine in thoracic epidural (10 and 30 microl) compared to high-volume (30 microl anaesthesia animals. Determination of the sympathetic blockade by means of laser Doppler perfusion imaging was restricted to the paws due to a higher density of subcutaneous blood vessels as compared to the abdominal wall (mean +/- SD: 3.93 +/- 0.06 vs. 1.35 +/- 0.05/384 mm(2), p < 0.05). Only high-volume (30 microl) lumbar and thoracic epidural anaesthesia (10 and 30 microl) increased skin perfusion in both hind and front paws. This extensive sympathetic blockade was demonstrated to include splanchnic segments using thermography. Segmental spread of methylene blue did not closely correspond to laser Doppler findings and should be interpreted as minimum rather than exact epidural spread of local anaesthetic.

Developing and implementing an interdisciplinary feeding training program within a large institution.

Planning is a daily part of every manager's job. Occupational therapy managers are frequently involved in the planning process and its component functions of designing, making decisions, delegating, controlling and evaluating. A11 of these management skills were called upon when a small occupational therapy department was assigned the program planning responsibility for developing a comprehensive feeding training program for a large institution.

The dog-ear rotation flap for the repair of large surgical defects on the head and neck.

BACKGROUND: The Mohs micrographic surgeon is often faced with the daunting challenge of having to repair very large surgical defects on the head and neck where cosmesis and maintenance of normal function are of paramount importance. OBJECTIVE: We describe a novel flap, the dog-ear rotation flap, for the repair of such defects. We will demonstrate that this flap offers superior cosmetic and functional results to many other closure options, particularly for extensive defects of the cheek, temple, forehead and scalp. METHODS: The dog-ear rotation flap is a combination repair. It is executed by first closing one end of the surgical defect in a primary side-to-side-fashion, to a point at which tension across the wound precludes any further closure. A rotation flap is then developed to close the remaining defect, using tissue from the large dog-ear created at the distal end of the wound. RESULTS: In our experience, the dog-ear rotation flap is able to close substantial head and neck defects with less tension across the wound edges when compared to other closure types, resulting in diminished scarring and little to no distortion of surrounding anatomic structures. It also provides an excellent tissue match, is relatively quick and easy to perform, and has an extremely low incidence of flap necrosis. CONCLUSIONS: The dog-ear rotation flap is an excellent choice for the repair of very large surgical defects on the head and neck, particularly the cheek, temple, forehead and scalp, and, in our experience, provides a superior cosmetic and functional result to other closure options.

Effect of honey bee venom on prostaglandin levels in mouse skin.

The effects of honey bee venom on prostaglandin (PG) E levels were studied in mouse skin under in vivo and in vitro conditions. Levels of PGE were increased 10.8-fold after 15 minutes exposure to reconstituted bee venom in vitro and 3.8-fold 35 minutes after a bee sting in vivo. Phospholipase A2 (PLA2), a major componet of bee venom, also caused a 10.9-fold increase in PGE levels in vitro and may be primarily responsible for this response of skin to bee venom.

Cerebral and subcutaneous cysticercosis treated with albendazole.

BACKGROUND: Cysticercosis is the most common parasitic disease of the central nervous system in the world, but cysticercosis cutis has been reported much less frequently. Because 54% of patients present with subcutaneous nodules, we report here the association of cysticercosis cutis in a patient with neurocysticercosis and review the literature and treatment options. CASE REPORT: The patient presented with multiple, asymptomatic subcutaneous nodules over the trunk and the extremities, associated with central nervous system involvement. Examination of an excised nodule by light microscopy revealed a larval cyst in the deep dermis surrounded partly by a fibrous pseudocapsule. Computed tomographic scanning af the skull showed multiple, nonenhancing, and calcified cycts in both cerebral hemispheres. Treatment with albendazole, 15 mg/kg/day for 30 days, was highly effective. At follow-up 6 months later, most subcutaneous nodules had disappeared or were markedly reduced in size, and the cerebral lesions had much improved. CONCLUSIONS: Albendazole, a newer paracidal drug, seems to be more effective and less expensive than some other drugs in use for the treatment of neurocysticercosis.

Purification and characterization of transforming growth factor-beta 2.3 and -beta 1.2 heterodimers from bovine bone.

A unique form of transforming growth factor-beta (TGF-beta), TGF-beta 2.3 heterodimer, has been purified from bovine bone extract. TGF-beta 2.3 migrated as a single 25-kDa band by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, whereas under reducing conditions it migrated as a 12.5 kDa band. The TGF-beta 2.3 reacted positively with anti-TGF-beta 2 and anti-TGF-beta 3 antibodies on immunoblots. Equal levels of TGF-beta 2 and TGF-beta 3 sequences were detected by N-terminal sequencing. TGF-beta 2.3 eluted as a single sharp peak by reverse-phase high performance liquid chromatography. However, prior reduction of the protein with dithiothreitol resulted in the protein eluting in two peaks, one containing predominantly TGF-beta 3 and the other containing predominantly TGF-beta 2. TGF-beta 2.3 inhibited proliferation of mink lung epithelial cells and promoted the formation of colonies of normal rat kidney fibroblasts in culture with specific biological activity similar to those of TGF-beta 1 and TGF-beta 2. These results demonstrate that the protein is TGF-beta 2.3 heterodimer, consisting of one polypeptide chain each of TGF-beta 2 and TGF-beta 3 linked by one or more disulfide bonds. In addition, TGF-beta 1.2 heterodimer, previously found only in porcine platelets, has also been purified from bovine bone extract.

Adrenergic regulation of sodium and chloride transport in the isolated cornea of rabbit and man.

Isolated corneas were mounted in Ussing-Zerahn-type chambers and short circuit current (SCC) was measured before and after application of drugs (5 X 10(-5) mol X 1(-1)) interfering with adrenergic receptors. Epinephrine increased SCC in the rabbit cornea and decreased SCC in the human cornea. alpha-Adrenergic stimulation or inhibition did not affect SCC. The increase in SCC observed after terbutaline (beta 2-agonist) was similar to the increase after isoproterenol (beta 1- and beta 2-agonist). SCC was not influenced by the beta 1-antagonist atenolol but was modified, although differently in rabbit and man, by the beta 1- and beta 2-antagonist propranolol. Thus, the catecholamine response of the rabbit and human cornea is mainly mediated by beta 2-adrenergic receptors. However, species differences were observed when testing the effect of propranolol on the transcorneal flux of 22Na and 36Cl. In the rabbit cornea the net Cl flux (directed from the aqueous to the tear side) was inhibited by propranolol, whereas net Na flux (from the tear to the aqueous side) was not influenced by the drug. In the human cornea propranolol reduced unidirectional Na flux from the aqueous to the tear side. Thus, the regulatory effect of propranolol on corneal transparency is different in man and the rabbit.

Long-term persistent infection of macaque monkeys with the simian immunodeficiency virus.

Juvenile rhesus macaques 6 to 18 months of age were experimentally infected by intravenous inoculation with the simian immunodeficiency virus (SIV), the T cell-tropic retrovirus of monkeys related to the human acquired immunodeficiency syndrome (AIDS) virus HIV. The SIV used for inoculation was grown either in normal human peripheral blood lymphocytes in the presence of interleukin 2 or in the human tumour cell line HUT-78. Eight of the macaques died 129 to 352 days post-inoculation with a variety of clinical and pathological findings paralleling those of AIDS in humans. However eight other animals became persistently infected for prolonged periods; these eight macaques remained alive at 537 and 820 days post-inoculation despite persistent lymphadenopathy and our continued ability to isolate SIV. The ability of these monkeys to survive infection correlated directly with the strength of their antibody response to SIV. Infection was also established in macaques using approximately 100 tissue culture infectious doses of HUT-78-grown SIV. There was no correlation between the dose of virus inoculum and either the strength of the antibody response or clinical outcome. These results demonstrate that SIV infection of macaques can be used not only to study acute AIDS but also to mimic the long-term persistent infection seen in carriers of HIV.

Simian models for AIDS.

The macaque immunodeficiency syndrome has many parallels to AIDS in humans. Affected monkeys develop profound, prolonged T lymphocyte dysfunction and die of lymphomas or opportunistic infections. We recently isolated a virus that we call SIV from four sick macaque monkeys. The morphology, growth characteristics, and antigenic properties of this virus indicate that it is related to the causative agent of human AIDS. The pathogenicity of this newly isolated virus was tested in macaque monkeys. Five of six died between 127 and 352 days following inoculation. The animals developed a wasting syndrome and died with adenovirus pancreatitis and/or pneumonia and primary retroviral encephalitis. Immunological abnormalities in these animals included a decrease in circulating T4+ lymphocytes and depressed peripheral blood lymphocyte proliferative response to pokeweed mitogen. The SIV monkey model holds great promise for testing antiviral agents and for the development of vaccines against AIDS.

Loss of cytomegalovirus-specific CD4+ T cell responses in human immunodeficiency virus type 1-infected patients with high CD4+ T cell counts and recurrent retinitis.

Clinical histories are reported for 2 patients treated with highly active antiretroviral therapy (HAART) who experienced multiple relapses of cytomegalovirus (CMV) retinitis, despite suppression of human immunodeficiency virus type 1 (HIV-1) viremia and improvement in CD4+ T cell counts (to >400 cells/microL). CMV-specific CD4+ T cell immune reconstitution was measured directly, using cytokine flow cytometry, which revealed persistent deficits in CMV-specific CD4+ T cell responses in both patients. CMV-specific T cells constituted 0.14% and 0.05% of the total CD4+ T cell count in these patients, which is significantly lower than the percentages for 34 control subjects (0.6%-46%; CD4+ T cell count range, 7-1039 cells/microL; P=.019). Deficits in pathogen-specific immune responses may persist in some individuals, despite suppression of HIV-1 replication and substantial increases in circulating CD4+ T cells after HAART, and such deficits may be associated with significant morbidity from opportunistic infections.

Direct measurement of CD4+ and CD8+ T-cell responses to CMV in HIV-1-infected subjects.

Data from murine models of chronic viral infection suggest that CD4+ T-cell responses to viral pathogens are important in sustaining the number and/or function of CD8+ cytotoxic T-cell (CTL) effectors. In this study, we used cytokine flow cytometry (CFC), staining with HLA-A*0201-peptide tetramers, and peptide stimulation with epitopic peptides to study functional CD4+ and CD8+ T-cell responses to cytomegalovirus (CMV) in human subjects coinfected with CMV and the human immunodeficiency virus, type 1 (HIV-1). We show that strong CD4+ and CD8+ T-cell responses to CMV antigens are sustained over time in HIV-1-infected individuals. Those who maintain a strong CD4+ T-cell response to CMV are also likely to maintain higher frequencies of CD8+ T cells capable of binding to HLA-A*0201-CMV pp65 (A2-pp65) tetramers as well as responses to pp65 peptide stimulation with effector cytokine production. These data support the hypothesis that declines in frequencies of CD4+ T-cell responses to CMV are associated with an inability to sustain high levels of CMV-specific CD8+ T-cell responses in HIV-1-infected subjects. These declines may precede the onset of CMV-associated end organ disease.

Study of long-term cultures of simian immunodeficiency virus (SIVmac 251)-infected peripheral blood lymphocytes.

A culture of rhesus monkey peripheral blood lymphocytes was divided into two parts; one was kept as an uninfected control, and the other was infected with a strain of simian immunodeficiency virus (SIVmac251) originally isolated from a rhesus monkey that died of a malignant lymphoma associated with acquired immune deficiency syndrome. Both cultures were sampled at successive intervals from 1 to 40 days postinfection. Each sample was subjected to in situ hybridization for detection of viral mRNA, immunocytochemical detection of viral core protein (p27), reverse transcriptase assay, electron microscopy, and immunophenotypic characterization of infected cells. These techniques were used to define viral growth kinetics of this novel lentivirus in peripheral blood lymphocytes. The first evidence of SIVmac251 replication was obtained by an in situ hybridization signal for viral mRNA at 2 days postinoculation. This was followed by detection of viral p27 core protein by immunocytochemistry on day 4. Reverse transcriptase activity above control values was not detected until day 8. Budding particles were not found in the infected cultures until 14 days postinfection. Results of in situ hybridization, immunocytochemistry, and reverse transcriptase assay indicated that two bursts of viral replication occurred during the course of this study. The first, at 3 weeks postinfection, was due to infection and subsequent depletion of CD4+ lymphocytes, while the second, 3 weeks later, resulted from a cycle of replication in CD8+ lymphocytes and the remaining CD4+ cells, culminating in the death of all cells on day 39 postinoculation.

Characterization of infectious molecular clones of simian immunodeficiency virus (SIVmac) and human immunodeficiency virus type 2: persistent infection of rhesus monkeys with molecularly cloned SIVmac.

Infection of macaque monkeys with simian immunodeficiency virus (SIV) is probably the best animal model currently available for studying acquired immunodeficiency syndrome. In this report, we describe three infectious molecular clones of SIVmac and one of human immunodeficiency virus type 2 (HIV-2) and their use in the study of cell and species specificity, animal infection, and the relationship of gene sequence to function. Replication of the cloned viruses in different cell lines varied dramatically. Some human CD4+ cell lines (HUT 78 and MT-4) supported the replication of SIVmac and HIV-2, while others (CEM and Jurkat-T) supported the replication of HIV-2 but not SIVmac. Growth of cloned virus in macaque lymphocytes in vitro was predictive of macaque infection in vivo. Macaque lymphocytes supported the replication of SIVmac239 and SIVmac251 but not SIVmac142 or HIV-2ROD. Using virus recovery and antibody response as criteria for infection, macaques that received cloned SIVmac251 and SIVmac239 became infected, while macaques receiving cloned SIVmac142 and HIV-2ROD did not become infected. Nucleotide sequences from the envelope region of all four cloned viruses demonstrated that there is considerable flexibility in the location of the translational termination (stop) signal. These infectious molecular clones will be very useful for future studies directed at the molecular basis for persistence, pathogenicity, tropism, and cell and species specificity.

Bovine bone activin enhances bone morphogenetic protein-induced ectopic bone formation.

A 25-kDa homodimeric protein was purified from demineralized bovine bone extract and identified as activin A. The bovine bone activin enhanced formation of ectopic bone in rat subcutis when implanted in combination with partially purified bovine bone morphogenetic protein (BMP-2, BMP-3) in collagen/ceramic carrier. The implants, removed at 14 days, contained markedly elevated levels of alkaline phosphatase activity. Histological examination revealed an extensive formation of woven bone with very little cartilage. In contrast, a combination of transforming growth factor-beta 2 and BMP promoted formation of bone with an abundance of cartilage. The implants with BMP alone exhibited some osteoinductive activity, while the implants with activin alone showed no activity. These results demonstrate that bone is a rich source of activin and that activin plays an important role in modulating bone formation.

Simian immunodeficiency virus from African green monkeys.

Simian immunodeficiency virus (SIV) was isolated from the total peripheral blood mononuclear cell population and the monocyte-macrophage adherent cell population of three seropositive green monkeys originating from Kenya. SIV from these African green monkeys (SIVagm) was isolated and continuously produced with the MOLT-4 clone 8 (M4C18) cell line but not with a variety of other cells including HUT-78, H9, CEM, MT-4, U937, and uncloned MOLT-4 cells. Once isolated, these SIVagm isolates were found to replicate efficiently in M4C18, SupT1, MT-4, U937, and Jurkat-T cells but much less efficiently if at all in HUT-78, H9, CEM, and MOLT-4 cells. The range of CD4+ cells fully permissive for replication of these SIVagm isolates thus differs markedly from that of previous SIV isolates from macaques (SIVmac). These SIVagm isolates had a morphogenesis and morphology like that of human immunodeficiency virus (HIV) and other SIV isolates. Antigens of SIVagm and SIVmac cross-reacted by comparative enzyme-linked immunosorbent assay only with reduced efficiency, and optimal results were obtained when homologous antibody and antigen were used. Western blotting (immunoblotting) of purified preparations of SIVagm isolate 385 (SIVagm385) revealed major viral proteins of 120, 27, and 16 kilodaltons (kDa). The presumed major core protein of 27 kDa cross-reacted antigenically with the corresponding proteins of SIVmac (28 kDa) and HIV-1 (24 kDa) by Western blotting. Hirt supernatant replicative-intermediate DNA prepared from cells freshly infected with SIVagm hybridized to SIVmac and HIV-2 DNA probes. Detection of cross-hybridizing DNA sequences, however, required very low stringency, and the restriction endonuclease fragmentation patterns of SIVagm were not similar to those of SIVmac and HIV-2. The nucleotide sequence of a portion of the pol gene of SIVagm385 revealed amino acid identities of 65% with SIVmac142, 64% with HIV-2ROD, and 56% with HIV-1BRU; SIVagm385 is thus related to but distinct from previously described primate lentiviruses SIVmac, HIV-1, and HIV-2. Precise information on the genetic makeup of these and other SIV isolates will possibly lead to better understanding of the history and evolution of these viruses and may provide insight into the origin of viruses that cause acquired immunodeficiency syndrome in humans.