Isolation and characterization of HSV-1 DNA from trigeminal ganglion neurons during suppressed infection in vitro.

The current study analyzed the herpes simplex virus type 1 (HSV1) genome during suppressed and reactivated infection in vitro. Utilization of 3H-labeled HSV1 provided a highly specific probe for intracellular localization, isolation, and characterization of the HSV genome after infection of rabbit trigeminal ganglion neurons. Restriction enzyme analysis of viral DNA extracted during both suppressed and reactivated infection matched that of the HSV1 control DNA. Viral DNAs extracted from both cellular and nuclear fractions of host cells exhibited identical patterns. No detectable alterations in terminal fragments were observed, which suggests that the HSV1 genome is both linear and nonintegrated during suppressed infection in this system.

Transcriptional activity of HIV-1 and HHV-6 in retinal lesions from AIDS patients.

This study determined the frequency of multiple viral (HIV-1, HHV-6, and CMV) infections in 26 retinas from 16 AIDS patients. Of the 12 retinas of 26 that tested positive for HIV-1 DNA sequences, seven also were positive for HHV-6 DNA sequences. Four of these seven retinas were culture positive for HIV-1 and two of the four contained CMV DNA sequences and antigens. Using RNA probes, HIV-1 and HHV-6 transcriptional activity was demonstrated in two of the four HIV-1 culture positive retinas. These retinas also contained CMV DNA sequences and antigens. The results demonstrate that more than 35% of AIDS patients suffer from at least two simultaneous viral infections and 15% suffer from three viral infections. The presence of transcriptional activity of HIV-1 and HHV-6 suggests an active infection.

Localization of 3H-thymidine-labeled HSV-1 in latently infected rabbit trigeminal ganglion cells.

Although current data favor conservation of virus in a nonreplicating form during latency, the actual host cell-virus relationship during this quiescent period remains an enigma. The purpose of this study was to develop a highly specific probe for direct localization of the HSV type 1 (HSV-1) genome in an animal model that closely mimics human disease. Tritium-labeled HSV-1 was inoculated onto trigeminal ganglion (TG) neurons in vitro and onto New Zealand white rabbit corneas in vivo. During acute infection in vivo and after establishment of latency in vivo or suppressed infection in vitro the TG neurons were processed for autoradiography. Silver grains were localized over nuclei of 8-10% of TG neuron cell bodies during suppressed infection in vitro. Acute infection in vivo resulted in the localization of label over 5-10% of neuron cell bodies and satellite cells per section. During latency the label appeared over nuclei of 1-10% of TG bodies per section. This study shows that directly labeled HSV-1 can be found in TG neuron nuclei both in vivo and in vitro. It also suggests that HSV genetic material is lost from certain neurons when latency is established.

On spinal osteochondromas.

Osteochondromas (or osteocartilaginous exostoses) make up about 30% to 40% of benign bone tumors. Most are solitary lesions but some are multiple, usually with autosomal dominant inheritance. From 1% to 4% of osteochondromas occur in the spine, where they can cause a variety of signs and symptoms, including those of spinal cord or spinal root compression. The authors present five patients with osteochondromas of the spine and review the findings together with those of over 130 cases reported since 1907. The cases were divided into: 1) spinal osteochondromas in patients with multiple osteochondromas, and 2) solitary osteochondromas occurring in the spine. The age (mean +/- standard error of the mean) of patients in the first group was 21.6 +/- 1.8 years compared to 30.0 +/- 2.1 years for those in the second group (p less than 0.02). There was a significant male predominance overall (M:F = 2.5:1; p less than 0.0005). In both groups, one-half of the lesions involved the cervical spine. Symptoms are caused by pressure on adjacent structures. Spinal cord compression was reported more than twice as frequently in the multiple osteochondroma group as in the single osteochondroma group (77% vs 33%; p less than 0.0005). Computerized tomography (CT) is the imaging procedure of choice. In both groups, the majority of surgically treated patients (90% and 88%, respectively) improve, with about three-quarters of the improved patients having no residual disease or only minor deficits.

Study of HSV-1 DNA species from trigeminal ganglia of rabbits during acute and latent infections.

Recurrent herpetic keratitis remains a major cause of corneal blindness in developed countries. A fundamental unanswered question regarding herpes simplex virus infection concerns the relationship between the virus and host cell DNA during latency. In the present study DNA was extracted from trigeminal ganglia during both acute and latent infection following ocular inoculation. Extracted, purified DNA was utilized for transfection and for hybridization studies using a 32P-labeled HSV-1 DNA probe. DNA extracted during acute infection was complete, linear and non-integrated. Autoradiographic patterns of DNA isolated during latent infection were suggestive of two separate DNA species.

Frequency of dual infections of corneas with HIV-1 and HHV-6.

In the current study, 35 pairs of corneas from asymptomatic carriers of HIV-1 and ten pairs from AIDS patients were analyzed for the presence of HIV-1 and HHV-6. The tissues were evaluated for viral antigens, transcripts, DNA sequences and intact and infectious virus. Three corneas from two asymptomatic carriers of HIV-1 and three corneas from two AIDS patients were culture positive for HIV-1. One of the three HIV-1 positive corneas from an asymptomatic HIV-1 carrier also was culture positive for HHV-6. Two of the tissue culture positive corneas from asymptomatic HIV-1 carriers and two from AIDS patients also tested positive for HIV-1 transcriptional activity by in situ hybridization. The label denoting the transcriptional activity was limited to stromal keratocytes. Most significantly, we were able to demonstrate the presence of HIV-1 particle(s) in sections and cultured PBMC from one of the HIV-1 culture positive corneas. PBMC from the same cornea also contained herpes virus particles. This report strengthens our earlier findings that HIV-1 and HHV-6 can invade corneal tissue, which emphasizes the importance of vigorous screening of corneal donors, specifically donors with HIV-1 exposure.

Demonstration of HIV-1 and HHV-6 in AIDS-associated retinitis.

Using an immunohistochemical technique and monoclonal antisera, HIV-1 and CMV antigens were demonstrated in lesioned areas of retinal tissues from selected AIDS patients. Polymerase chain reaction (PCR) was utilized to detect HIV-1 and HHV-6 DNA sequences in total retinal tissues from these patients. In this study of six eyes from four patients, two of the retinas contained three different viruses, HIV-1, HHV-6 and CMV. To determine whether HIV-1 and HHV-6 DNA sequences were restricted to the intraretinal lesions, normal and lesioned areas were dissected from the retina, DNA was extracted and subjected to amplification using PCR. The results showed that HIV-1 and HHV-6 DNA were restricted to the lesioned areas. All four lesions (from two different patients) utilized in this study showed the presence of CMV antigens immunohistochemically. The combination of viruses present in each lesion was either HIV-1 and CMV or HHV-6 and CMV. Two of four lesions contained HIV-1 and CMV; a third lesion showed the presence of HHV-6 and CMV. The fourth lesion contained only CMV antigens.

The incidence of HIV-1 and HHV-6 in corneal buttons.

Twenty pairs of corneas from asymptomatic carriers of HIV-1 and seven pairs from AIDS patients were analyzed for the presence of HIV-1 and HHV-6 antigens, viral transcripts, DNA sequences, and intact and infectious particles. Although serum from all donors was positive for both HIV-1 and HHV-6 antibody by Western blot analysis, only one cornea from an asymptomatic carrier of HIV-1 was positive for HIV-1 and HHV-6. The cornea was positive when tested by tissue culture, PCR, in situ hybridization, and electron microscopy. There was no tear film contamination. These results suggest that HIV-1 and HHV-6 may be capable of invading corneal tissue.

Immunoglobulin heavy chain rearrangements in primary brain lymphomas. A study using PCR to amplify CDR-III.

Primary brain lymphomas (PBLs) have only rarely been analysed for immunoglobulin heavy chain (IgH) rearrangements. In this study, DNA was extracted from paraffin blocks in 23 cases of PBL and examined for IgH rearrangements using the polymerase chain reaction (PCR) to amplify the complementarity-determining region III (CDR-III) of rearranged IgH genes. Fifteen of the cases were phenotyped on paraffin-embedded tissue using a pan-B and pan-T antibody (L26 and UCHL-1, respectively). The remaining eight cases were not phenotyped for lack of tissue. For comparison, we used DNA extracted from paraffin blocks of normal brain, lymph nodes with lymphoid hyperplasia, and non-lymphoid malignancies. PCR products were examined by polyacrylamide gel electrophoresis. Among the ten B-cell PBL; four had a pattern indicative of IgH rearrangement, one had a germline pattern, and five had no detectable PCR products. Among the five T-cell PBLs, one had a germline pattern and four had no detectable products. Among the eight untyped PBLs, two had IgH rearrangement, four had a germline pattern, and two gave no detectable products. DNA from non-lymphoid tissues gave a consistent germline pattern, while DNA from polyclonal lymphoid populations (lymph node) had a pattern of polyclonal IgH rearrangement. In a dilution study, a clonal rearrangement could be detected as long as the clone's DNA constituted at least 10 per cent of the total DNA. PCR to amplify CDR-III can be successfully applied to DNA extracted from paraffin blocks, and it detected a clonal rearrangement in 50 per cent of cases that gave a detectable pattern.(ABSTRACT TRUNCATED AT 250 WORDS)

Reconstitution of the patellar tendon donor site after graft harvest.

Little is known about the fate of the donor site after the central 1/3 of the patellar tendon is harvested for anterior cruciate ligament reconstruction. This study evaluated the donor site in the patellar tendon at various times after a graft from the central 1/3 of the patellar tendon was harvested. Fourteen patients were studied with magnetic resonance images taken from 6 weeks to 2 years after anterior cruciate ligament reconstruction. Axial, sagittal, and coronal views of the patellar tendon were obtained. A second group of 8 patients who had previous anterior cruciate ligament reconstruction were returned to the operating room for subsequent procedures on the affected knee. These procedures were performed 2 to 24 months after the original reconstruction. An open biopsy was obtained from the donor site in the patellar tendon. On magnetic resonance images, the size of the defect and the intensity of the signal in the central 1/3 of the tendon decreased with time from surgery. At 2 years, the defect was indistinguishable from normal tendon. Histologically, the scar in the defect progressively matured with time, becoming nearly identical to normal tendon at 2 years.