Mouse cytomegalovirus reactivation in severe combined immune deficient mice after implantation of latently infected salivary gland.

The hypothesis that replication-competent mouse cytomegalovirus (MCMV) is detectable in severe combined immunodeficient (scid) mice after implantation of latently infected tissue was examined. Sections of latently infected salivary gland from 5 BALB/c mice were implanted into 20 C.B-17 scid mice. Recipient scid mice were sacrificed weekly for 5 weeks, and MCMV infection was detected in target organs using culture and DNA polymerase chain reaction (PCR). All donors were negative by standard culture but positive by DNA PCR. Replicating MCMV was recovered from 9 of 15 recipient scid mouse salivary gland, lung, liver, or spleen samples at postoperative weeks 2-5. No virus was recovered from recipient scid mice at weeks 1 or 12. Transplantation of latently infected tissues into scid mice resulted in rapid reactivation and dissemination of the virus. Further study of this model promises insight into the mechanisms of CMV latency and reactivation.

Experimental murine cytomegalovirus infection in severe combined immunodeficient mice.

Severe combined immunodeficient (scid) and BALB/c mice were experimentally infected with murine cytomegalovirus (MCMV). Scid mice infected by the intraperitoneal route died or were moribund at dose-dependent times ranging from 12 to 13 days after inoculation for > or = 3.00 x 10(4) plaque forming units (pfu) of virus to 25 days for 1.17 x 10(2) pfu. Histologic lesions included severe adrenal necrosis at low doses and splenic necrosis at high doses. Multinucleate hepatocytes with multiple inclusion bodies were observed at all doses. In visceral organs, the inflammatory response consisted of cell necrosis and neutrophil infiltration. Scid mice infected with 1.00 x 10(3) pfu by the intranasal route were moribund by 24 or 25 days after inoculation. Viral titers in tissues examined from these mice increased in all organs examined until they became moribund. BALB/c mice infected intranasally had detectable virus titers in the adrenal glands, salivary glands, lungs, and spleen by 7 or 14 days after inoculation, but decreased thereafter. These mice remained clinically normal through the infection. In BALB/c mice, histologic lesions were present only in the salivary glands.

Pulsatile reperfusion after cardiac arrest improves neurologic outcome.

Cardiopulmonary bypass (CPB) using nonpulsatile flow (NPF) is advocated for refractory cardiac arrest. This study examined cerebral outcome after resuscitation with pulsatile flow (PF) versus NPF. Dogs arrested for 12.5 minute were reperfused with NPF (n = 11) using roller pump CPB or PF (n = 11) using mechanical biventricular cardiac massage. Pump flows were similar between groups; however early arterial pressures were greater during PF versus NPF, *p less than 0.05. Circulatory support was weaned at 60 minutes' reperfusion. Neurologic recovery of survivors (n = 16) was significantly better after PF versus NPF, *p = 0.01. The presence of brain lesions on magnetic resonance images did not significantly differ between groups at 7 days. Brain then were removed and regions examined for ischemic changes. Loss of CA1 pyramidal neurons was more severe after NPF versus PF, +p = 0.009. Ischemic changes were more frequent after NPF in the caudate nucleus (+p = 0.009) and watershed regions of the cerebral cortex (+p = 0.062), compared with PF. These results demonstrate that PF improves cerebral resuscitation when treating cardiac arrest with mechanical circulatory support (* = MANOVA with repeated measures, + = categorical data analysis.

Murine cytomegalovirus gene amplification and culture after submaxillary salivary gland biopsy.

An important target tissue for murine cytomegalovirus (CMV) infection is the submaxillary salivary gland. Submaxillary salivary gland biopsy specimens from BALB/c mice latently infected with murine CMV were examined for murine CMV DNA by in vitro enzymatic amplification using the polymerase chain reaction preceding oligonucleotide hybridization. The amplified sequence was a 152-base pair segment from within the immediate early gene of murine CMV. Biopsy and whole gland specimens from acutely infected BALB/c mice and latently infected, immunosuppressed BALB/c mice were compared for active murine CMV infection. After acute infection with murine CMV, virus was recovered in all cultures of both biopsy and whole salivary gland specimens but from none of the latently infected animals. Reactivated virus was detected by culture of both biopsy (90%) and whole salivary gland specimens (100%) from latently infected mice that received antithymocyte serum. Viral nucleic acid was detected in 90% of biopsy specimens from latently infected animals. Hence, active murine CMV infection can be detected in biopsy specimens from mice with acute and reactivated infection and murine CMV DNA can be amplified and detected in salivary gland biopsy specimens from latently infected animals. Biopsy of this or other target tissues can be useful for obtaining tissue for viral studies where the survival of the animal is important and it is useful to distinguish latent from acute or reactivated infection.

Cell cycle-dependent initiation of hepatocarcinogenesis in rats by (+/-)-7r,8t-dihydroxy-9t,10t-epoxy-7,8,9,10-tetrahydrobenzo(a)py rene.

Male F344 rats were treated once with (+/-)-7r,8t-dihydroxy-9t,10t-epoxy-7,8,9,10-tetrahydrobenzo(a)pyre ne (BPDE) at various times after a two-thirds partial hepatectomy. Diet containing 0.05% phenobarbital was fed subsequently to promote the development of hepatocellular neoplasms from initiated cells. This combined treatment caused an apparently continuous emergence of hepatocellular neoplasms between 31 and 60 weeks after initiation. Yields of hepatocellular neoplasms (adenomas and carcinomas) seen at 45 weeks after initiation varied according to the time of treatment with BPDE. Rats treated at 15 h after partial hepatectomy, when maximal fractions of proliferating hepatocytes were entering the S phase, displayed the greatest incidence and yield of hepatocellular neoplasms. Rats treated when hepatocytes were early in the prereplicative G1 phase demonstrated a significantly lower incidence and yield of hepatocellular neoplasms. No neoplasms were seen in livers of rats treated with BPDE without prior partial hepatectomy; this indicates the insensitivity or resistance of nonproliferating G0 hepatocytes to initiation by this chemical. The variation in yields of neoplasms could not be attributed to variation in binding of carcinogen to DNA or to rates of DNA repair. The risk of initiation of hepatocytes appeared to be correlated (r = 0.95) with the fraction of hepatocytes that were entering the S phase at the time of treatment with BPDE. These results reveal a significant cell cycle dependency for initiation of hepatocarcinogenesis by BPDE and show that proliferating hepatocytes are at greatest risk when early in the S phase. Quantitation of islands of hepatocellular alteration suggested a similar pattern of cell cycle-dependent sensitivity, although the cycle-dependent variation in island frequencies (1.4-fold) was less than the variation in yields of neoplasms (5-fold). However, 1300-5000 islands were estimated to appear for every one neoplasm that appeared. When viewed within the context of multistep carcinogenesis, and assuming that islands of altered hepatocytes represent a step in this process, then these results suggest that islands may represent populations of cells at least one step removed from neoplasia.

Quantitative relationship between initiation of hepatocarcinogenesis and induction of altered cell islands.

We have quantified the initiation of hepatocytic neoplasms and the induction of altered cell islands in regenerating livers of rats given a single treatment with one of three carcinogens before or during the peak of DNA synthesis after partial hepatectomy. For up to 20 wk after treating livers during the peak of DNA synthesis with methyl(acetoxymethyl)nitrosamine (DMN-Ac), hepatocytic neoplasms were not seen. Thereafter, in rats fed the liver tumor promoter, phenobarbital, neoplasms emerged continuously so that by 60 wk after initiation, livers held an average of 5.5 neoplasms. Islands of cellular alteration, identified by their abnormal retention of glycogen on fasting, also appeared to emerge continuously between 20 and 60 wk after initiation. By 60 wk, promoted livers contained about 10,000 islands. In DMN-Ac-initiated, phenobarbital-promoted livers, neoplasms and islands maintained a constant numerical relationship over time with about 1,450 islands emerging for every neoplasm that emerged. This ratio of islands to neoplasms differed according to the type of carcinogen used to initiate hepatocarcinogenesis and depending on whether promotion with phenobarbital was included. In livers initiated with DMN-Ac but not promoted with phenobarbital, the ratio of islands to neoplasms was about 7,750:1. In livers initiated by treatment with (+/-)-7 alpha, 8 beta-dihydroxy-9 beta, 10 beta-epoxy-7,8,9, 10-tetrahydrobenzo[alpha]pyrene at the peak of DNA synthesis and then promoted with phenobarbital, the ratio of islands to neoplasms was 7,200:1. In livers exposed to gamma rays at the peak of DNA synthesis in regenerating livers and promoted, no neoplasms were seen in our sample although islands could be enumerated.(ABSTRACT TRUNCATED AT 250 WORDS)