'Value-based methodology for person-centred, integrated care supported by Information and Communication Technologies' (ValueCare) for older people in Europe: study protocol for a pre-post controlled trial.

BACKGROUND: Older people receive care from multiple providers which often results in a lack of coordination. The Information and Communication Technology (ICT) enabled value-based methodology for integrated care (ValueCare) project aims to develop and implement efficient outcome-based, integrated health and social care for older people with multimorbidity, and/or frailty, and/or mild to moderate cognitive impairment in seven sites (Athens, Greece; Coimbra, Portugal; Cork/Kerry, Ireland; Rijeka, Croatia; Rotterdam, the Netherlands; Treviso, Italy; and Valencia, Spain). We will evaluate the implementation and the outcomes of the ValueCare approach. This paper presents the study protocol of the ValueCare project; a protocol for a pre-post controlled study in seven large-scale sites in Europe over the period between 2021 and 2023. METHODS: A pre-post controlled study design including three time points (baseline, post-intervention after 12 months, and follow-up after 18 months) and two groups (intervention and control group) will be utilised. In each site, (net) 240 older people (120 in the intervention group and 120 in the control group), 50-70 informal caregivers (e.g. relatives, friends), and 30-40 health and social care practitioners will be invited to participate and provide informed consent. Self-reported outcomes will be measured in multiple domains; for older people: health, wellbeing, quality of life, lifestyle behaviour, and health and social care use; for informal caregivers and health and social care practitioners: wellbeing, perceived burden and (job) satisfaction. In addition, implementation outcomes will be measured in terms of acceptability, appropriateness, feasibility, fidelity, and costs. To evaluate differences in outcomes between the intervention and control group (multilevel) logistic and linear regression analyses will be used. Qualitative analysis will be performed on the focus group data. DISCUSSION: This study will provide new insights into the feasibility and effectiveness of a value-based methodology for integrated care supported by ICT for older people, their informal caregivers, and health and social care practitioners in seven different European settings. TRIAL REGISTRATION: ISRCTN registry number is 25089186 . Date of trial registration is 16/11/2021.

Cell-mediated cytotoxicity of murine cytomegalovirus-infected target cells allows for release of residual infectious virus.

Although cell-mediated cytotoxicity effectively kills target virus-infected cells, no careful consideration has been given to the fate of infectious progeny virus contained within target cells following the cytolytic event. To address this issue with respect to murine cytomegalovirus (MCMV) pathogenesis, we developed a rapid semiquantitative assay for infectious MCMV based on expression of beta-galactosidase using a LacZ-expressing recombinant MCMV. Simultaneous use of this assay in combination with a modified cell-mediated cytotoxicity assay revealed that cytotoxicity of MCMV-infected target cells mediated by MCMV-immune splenic cells does not lead to inactivation of intracellular infectious virus.

Perforin knockout mice, but not mice with MAIDS, show protection against experimental cytomegalovirus retinitis after adoptive transfer of immune cells with a functional perforin cytotoxic pathway.

Adoptive transfer studies were performed to test the hypothesis that the perforin cytotoxic pathway is more important than the Fas/FasL cytotoxic pathway in protection against experimental murine cytomegalovirus (MCMV) retinitis. Splenic immune cells from donor MCMV-immunized normal mice or gld mice deficient in Fas/FasL-mediated cytotoxicity significantly reduced the frequency and severity of MCMV retinitis following subretinal MCMV challenge when transferred into recipient PKO mice deficient in perforin-mediated cytotoxicity. In sharp contrast, splenic cells from donor MCMV-immunized PKO mice failed to provide protection against MCMV retinitis when transferred into recipient PKO mice. Protection was not achieved, however, in recipient mice with retrovirus-induced immunodeficiency (MAIDS), even when splenic cells originated from MCMV-immunized normal mice.

The prevalence of insulin autoantibodies at the onset of Type 1 diabetes is higher in males than females during adolescence.

AIMS/HYPOTHESIS: The incidence of Type 1 diabetes shows little sex bias up to age 15 years, but more males are diagnosed in early adult life. Humoral responses to the beta cell antigen insulin could help to reveal the mechanism underlying this difference. We therefore determined the influence of sex on the prevalence of insulin autoantibodies (IAA) at diagnosis. METHODS: IAA were measured by radiobinding assay in 598 patients with newly diagnosed Type 1 diabetes (aged 10.5, range 0.8-20.7 years, 333 male), and analysed according to age, sex and HLA class II genotype. RESULTS: Overall, 74% of males and 65% of females had IAA above the 97.5(th) centile of 2860 schoolchildren ( p=0.028). IAA prevalence was similar in males and females under the age of 15 (0-4 yr, 95% vs 88%; 5-9 yr, 76% vs 73%; 10-14 yr, 67% vs 58%), but male excess was seen between 15 and 21 years (66% vs. 32%, p(corr)=0.016). HLA class II genotype was available for 426 patients. IAA prevalence in DR4 homozygous patients was 87%, in DR4 heterozygous patients 72% and in DR4 negative patients 55% ( p<0.001). Multivariate analysis showed independent association of IAA with age ( p<0.001), number of DR4 alleles ( p<0.001) and male sex ( p=0.002). CONCLUSIONS/INTERPRETATION: The prevalence of IAA in patients with newly diagnosed Type 1 diabetes is higher in males than females between 15 and 21 years of age. The lower prevalence of IAA in adolescent females implies sex-specific modulation of the autoimmune process during puberty.

Three generations of autoimmune diabetes: an extended family study.

AIMS/HYPOTHESIS: One in four children with Type 1 diabetes in a population-based family study has an affected grandparent. We set out to study the clinical and immune features of diabetes in the grandparents' generation, and to examine sharing of HLA class II susceptibility haplotypes between grandparent and grandchild. METHODS: Of 5855 grandparents in the Bart's-Oxford family study, 428 (7.3%) were known to have diabetes. Clinical data and samples were collected from 115 of 213 surviving affected grandparents and from 219 unaffected grandparents within the same families. Samples were tested for ICA and autoantibodies to GAD and IA-2, and typed for HLA-DRB1-DQA1-DQB1. Transmission of HLA class II haplotype from affected and unaffected grandparents to the diabetic proband was compared. RESULTS: Of 115 affected grandparents studied, the median age at diagnosis was 61 years and at analysis was 73 years; 70% were diet or tablet treated and 30% were on insulin. One or more islet autoantibodies were found in 26% and 66% had one or both of the high risk HLA class II susceptibility haplotypes DRB1*03-DQA1*0501-DQB1*0201 or DRB1*04-DQA1*0301-DQB1*0302. In 79 informative families the HLA class II haplotype of the affected grandparent was transmitted to the proband more frequently than expected overall (59%, p=0.02), and in the insulin-treated subgroups (65%, p=0.03). CONCLUSION/INTERPRETATION: A total of 7.3% of grandparents reported a clinical diagnosis of diabetes and 2.2% had features of Type 1 diabetes. Genetic susceptibility was shared between grandparents with diabetes and their affected grandchildren. Diabetes in the grandparents of children with Type 1 diabetes often has an autoimmune basis, even when it presents late in life and does not require insulin treatment.

Comparison of three approaches to doxorubicin therapy: free doxorubicin, liposomal doxorubicin, and beta-glucuronidase-activated prodrug (HMR 1826).

BACKGROUND: Three approaches to doxorubicin therapy are directly compared: free doxorubicin, liposomal doxorubicin and beta-glucuronidase-activated prodrug (HMR 1826). MATERIALS AND METHODS: The optimal dose of HMR 1826 was determined to be 200 mg/kg once a week and subsequent studies were carried out comparing HMR 1826 at 200 mg/kg 1x/wk, liposomal doxorubicin (Doxil) at 9 mg/kg 1x/wk and free doxorubicin at 7 mg/kg 1x/wk in seven different human tumor xenograft models. RESULTS: All three forms of doxorubicin inhibited tumor growth with similar efficacy in each of the tumor models with the exception of MDA-MB-231 tumor xenografts, which were resistant to free doxorubicin but sensitive to Doxil and HMR 1826. Overall less weight loss was observed with HMR 1826 treatment. CONCLUSIONS: The efficacy of HMR 1826 is equal to or better than that of doxorubicin and Doxil at a safe dose and schedule, indicating that the beta-glucuronidase activated prodrug approach is safe and effective.

Preclinical evaluation of the cardiac toxicity of HMR-1826, a novel prodrug of doxorubicin.

Cardiotoxicity represents the major side-effect limiting the clinical use of anthracyclines, especially doxorubicin, in cancer chemotherapy. The use of non-toxic prodrugs, or of liposome-encapsulated drugs, allows a better targeting of the tumours and may, therefore, improve the tolerance to the treatment. Using the model of isolated perfused rat heart, we have evaluated the cardiotoxicity of a novel prodrug of doxorubicin, HMR-1826, which consists of the association of doxorubicin to glucuronic acid. We have compared the cardiac effects (developed pressure, contractility and relaxation of the left ventricle) induced by HMR-1826 to those induced by doxorubicin and Doxil, a liposomal form of doxorubicin. HMR-1826 was administered intravenously every other day for 11 days at doses of 50-200 mg kg(-1) per injection while doxorubicin was administered according to the same protocol at doses of 1-3 mg kg(-1) per injection. Doxorubicin strongly decreased the cardiac functional parameters at the doses of 2.5 and 3 mg kg(-1) per injection. Doxil (3 mg kg(-1) and HMR-1826 (50-150 mg kg(-1)) were largely devoid of cardiotoxicity. HMR-1826 only induced significant alterations of the cardiac function at the highest dose used (200 mg kg(-1) per injection). These alterations were much lower than those of doxorubicin at 2.5 mg kg(-1) per injection, despite similar general toxicity symptoms (weight loss, nose bleeding and diarrhoea) at these respective doses. Thus, HMR-1826 appeared about 100-fold less cardiotoxic than doxorubicin.

Systemic murine cytomegalovirus infection of mice with retrovirus-induced immunodeficiency results in ocular infection but not retinitis.

Experiments were performed to explore the ability of murine cytomegalovirus (MCMV) to disseminate to the eye following intravenous inoculation and to cause infection of ocular tissues and necrotizing retinitis in C57BL/6 mice with retrovirus-induced immunodeficiency syndrome (MAIDS). Although infectious virus could be detected in whole eye homogenates of mice with MAIDS at 10 days after intravenous MCMV inoculation, a recombinant MCMV (RM461) that carries an MCMV IE1 promoter-LacZ insert was used as a tracer virus to confirm direct infection of ocular tissues. Evidence for MCMV replication (determined by RM461-induced expression of beta-galactosidase) was consistently observed in the nonpigmented epithelium of the ciliary body of the eyes of MAIDS animals at 14 days after infection. In sharp contrast, however, the neurosensory retina was spared and necrotizing retinitis failed to develop. These findings demonstrate that systemic MCMV infection of mice with MAIDS results in ocular MCMV infection without development of ocular MCMV disease. Conversion of occult subclinical MCMV infection of ocular tissues to overt clinical MCMV retinitis may require as yet unidentified cofactor(s). Identification of these cofactors could lead to more innovative therapeutic approaches for prevention and/or treatment of CMV retinitis in patients with AIDS.

Penetration of levofloxacin into lung tissue after oral administration to subjects undergoing lung biopsy or lobectomy.

STUDY OBJECTIVE: To evaluate the pulmonary tissue distribution of levofloxacin, the new once-daily fluoroquinolone, after a single 500-mg oral dose. DESIGN: Open-label study. SETTING: One pulmonary clinic and two university-affiliated teaching hospitals. PATIENTS: Eighteen adults undergoing lung biopsy or lobectomy. INTERVENTIONS: Levofloxacin plasma and lung tissue concentrations were determined by high-performance liquid chromatography with fluorescence detection. Lung tissue levofloxacin concentrations were corrected for blood contamination by measuring hemoglobin. MEASUREMENTS AND MAIN RESULTS: After a single 500-mg oral dose, concentrations of levofloxacin in lung tissue consistently exceeded those in plasma at every time point over the 24-hour sampling period, with tissue:plasma penetration ratios of 2.02 (2-3 hrs), 5.02 (4-6 hrs), 5.13 (11-17 hrs), and 4.13 (22-25 hrs). The mean penetration ratio over the 24-hour sampling period was 3.95 (range 1.06-9.98). Lung tissue concentrations of levofloxacin also exceeded minimum inhibitory concentration values for most community-acquired respiratory tract pathogens over the 24 hours. CONCLUSION: This study supports clinical evaluation of levofloxacin as once-daily oral therapy for community-acquired lower respiratory tract infections.

Effects of food and sucralfate on a single oral dose of 500 milligrams of levofloxacin in healthy subjects.

The effects of food and sucralfate on the pharmacokinetics of levofloxacin following the administration of a single 500-mg oral dose were investigated in a randomized, three-way crossover study with young healthy subjects (12 males and 12 females). Levofloxacin was administered under three conditions: fasting, fed (immediately after a standardized high-fat breakfast), and fasting with sucralfate given 2 h following the administration of levofloxacin. The concentrations of levofloxacin in plasma and urine were determined by high-pressure liquid chromatography. By noncompartmental methods, the maximum concentration of drug in serum (Cmax), the time to Cmax (Tmax), the area under the concentration-time curve (AUC), half-life (t1/2), clearance (CL/F), renal clearance (CLR), and cumulative amount of levofloxacin in urine (Ae) were estimated. The individual profiles of the drug concentration in plasma showed little difference among the three treatments. The only consistent effect of the coadministration of levofloxacin with a high-fat meal for most subjects was that levofloxacin absorption was delayed and Cmax was slightly reduced (Tmax, 1.0 and 2.0 h for fasting and fed conditions, respectively [P = 0.002]; Cmax, 5.9 +/- 1.3 and 5.1 +/- 0.9 microg/ml [90% confidence interval = 0.79 to 0.94] for fasting and fed conditions, respectively). Sucralfate, which was administered 2 h after the administration of levofloxacin, appeared to have no effect on levofloxacin's disposition compared with that under the fasting condition. Mean values of Cmax and AUC from time zero to infinity were 6.7 +/- 3.2 microg/ml and 47.9 +/- 8.4 microg x h/ml, respectively, following the administration of sucralfate compared to values of 5.9 +/- 1.3 microg/ml and 50.5 +/- 8.1 microg x h/ml, respectively, under fasting conditions. The mean t1/2, CL/F, CLR, and Ae values were similar among all three treatment groups. In conclusion, the absorption of levofloxacin was slightly delayed by food, although the overall bioavailability of levofloxacin following a high-fat meal was not altered. Finally, sucralfate did not alter the disposition of levofloxacin when sucralfate was given 2 h after the administration of the antibacterial agent, thus preventing a potential drug-drug interaction.

Systemic cytokine immunotherapy for experimental cytomegalovirus retinitis in mice with retrovirus-induced immunodeficiency.

PURPOSE: To evaluate and compare the in vivo administration of interleukin-2 (IL-2) or interleukin-12 (IL-12) in the immunotherapy of necrotizing retinitis caused by murine cytomegalovirus (MCMV) in mice with a retrovirus-induced immunodeficiency syndrome (MAIDS). METHODS: Adult C57BL/6 mice with MAIDS of 8 weeks' duration were treated with either a single intramuscular injection of polyethylene glycol-modified human recombinant IL-2 (PEG-IL-2) or multiple intramuscular injections of murine recombinant IL-12; untreated mice with MAIDS received phosphate-buffered saline. Two days later, the left eyes of all mice were inoculated with MCMV by subretinal injection and evaluated at day 6 for intraocular MCMV titers or at day 10 for frequency of necrotizing MCMV retinitis. RESULTS: Infectious MCMV was significantly reduced in whole eyes of PEG-IL-2-treated mice with MAIDS (2.8 log10), but not in whole eyes of IL-12-treated animals (4.4 log10) when compared with whole eyes of untreated animals with MAIDS (4.5 log10). Similarly, whereas eyes from approximately 80% of IL-12-treated and untreated mice with MAIDS showed histopathologic features consistent with classic necrotizing MCMV retinitis (full-thickness retinal necrosis associated with virus inclusions and cytomegalocytes), none (0%) of PEG-IL-2-treated animals with MAIDS showed classic MCMV retinitis. Instead, eyes from these animals showed either retinal folding or outer retinal atrophy, a pattern of histopathology similar to that observed in eyes from immunologically normal C57BL/6 mice inoculated subretinally with MCMV. CONCLUSIONS: These results provide proof-of-principle for the hypothesis that systemic cytokine immunotherapy will reduce the frequency of CMV retinitis in a setting of retrovirus-induced immunosuppression. Because of the striking differential effects of IL-2 and IL-12 on MCMV-retinitis in mice with MAIDS, the authors conclude that cytokine immunotherapy for cytomegalovirus-induced retinitis is cytokine-specific, even for such cytokines as IL-2 and IL-12 that have T cell regulation in common.

Antibody alone does not prevent experimental cytomegalovirus retinitis in mice with retrovirus-induced immunodeficiency (MAIDS).

Passive-transfer studies were performed to assess the ability of antibody alone to reduce the frequency and/or severity of necrotizing retinitis caused by murine cytomegalovirus (MCMV) in C57BL/6 mice with retrovirus-induced immunodeficiency syndrome (MAIDS). Initial experiments showed a gradual decline in the ability of mice to initiate humoral immunity during the evolution of MAIDS so that neither MCMV-specific IgM nor IgG could be detected during late-stage MAIDS. Passively administered hyperimmune MCMV immunoglobulin, however, could be detected within the serum of mice with MAIDS for at least 9 days after intraperitoneal injection and protected these animals in preliminary experiments from systemic MCMV disease and death when administered 24 h prior to intraperitoneal challenge with a lethal dose of virus. Nonetheless, passive transfer of hyperimmune MCMV serum to mice with MAIDS failed to reduce intraocular MCMV titers, frequency of retinitis, or severity of retinitis when administered 24 h prior to subretinal MCMV inoculation. Whereas whole eyes of MAIDS animals that received normal mouse serum and were injected subretinally with MCMV had an ocular MCMV titer of 4.3 log10 and a frequency of retinitis of 89% (severity score = 55%), whole eyes of antibody-treated mice with MAIDS had an ocular MCMV titer of 4.3 log10 and a frequency of retinitis of 87% (severity score = 57 %). Passive transfer of a neutralizing MCMV-specific monoclonal antibody also failed to reduce the frequency or severity of MCMV retinitis when administered to mice with MAIDS prior to subretinal MCMV inoculation. Our findings suggest that antibody immunotherapy alone will not be effective therapeutically for cytomegalovirus retinitis in patients with AIDS.

Mice immunosuppressed by murine retrovirus infection (MAIDS) are susceptible to cytomegalovirus retinitis.

Although various methods of immunosuppression have been used to enhance susceptibility of mice to murine cytomegalovirus (MCMV) retinitis, none reproduce the unique complexity of immune deficiency experienced by patients during the progression of AIDS. C57BL/6 mice are susceptible to a retrovirus-induced murine acquired immune deficiency syndrome (MAIDS), characterized by progressive immune dysfunction which shares many features with AIDS. We therefore evaluated the frequency and severity of MCMV retinitis in C57BL/6 mice with MAIDS. Following subretinal inoculation of MCMV, nearly 90% of mice with MAIDS developed a necrotizing retinitis 8 to 10 days postinfection, whereas retinitis was observed in only 8% of age-matched immunocompetent control mice. Histopathologic analysis of the retinitis that developed in mice with MAIDS revealed features similar to those found in AIDS-related CMV retinitis. Eyes from MAIDS animals also contained on average higher amounts of infectious virus when compared with eyes from control animals. We conclude that retrovirus-induced immunosuppression increases susceptibility of C57BL/6 mice to MCMV retinitis.

Diagnostic value for culture of cerebrospinal fluid from HIV-1-infected individuals for opportunistic viruses: a prospective study.

OBJECTIVE: To investigate the diagnostic value of cerebrospinal fluid (CSF) culture for opportunistic viruses from HIV-1-infected individuals. METHODS: A 4-year prospective study was conducted using a participant population consisting of 186 HIV-1-infected individuals without neurologic disease, 73 HIV-1-infected individuals with encephalopathy, myelopathy, and/or peripheral neuropathy, and 10 controls. CSF samples recovered at 1-year intervals were subjected to virus culture using technique commonly used in the clinical laboratory setting. RESULTS: CSF samples obtained from only 15 of the 269 (5.6%) participants yielded an opportunistic virus upon culture. Cytomegalovirus, herpes simplex virus types 1 and 2, adenovirus, and presumptive enteroviruses were identified. No consistent correlation was observed between the detection of an opportunistic virus within a CSF sample and the presence or future development of neurologic disease. However, a significant correlation was observed between culture of virus from CSF and the future development of abnormal CD4+ (chi 2, P = 0.0286) and CD8+ (chi 2, P = 0.0018) lymphocyte counts in HIV-1-infected participants without neurologic disease. CONCLUSION: These results show that culture of CSF to screen for opportunistic viruses is neither diagnostic nor predictive of neurologic disease in HIV-1-infected individuals. Nevertheless, the presence of virus within CSF may be an indicator of HIV-1-mediated immune dysfunction and a predictor for future development of abnormal CD4+ and/or CD8+ lymphocyte counts.

Staining characteristics and antiviral activity of sulforhodamine B and lissamine green B.

PURPOSE: Fluorescein and rose bengal are dyes used routinely in the examination of the ocular surface. As part of an ongoing search for a superior ophthalmic dye with optimal specificity and sensitivity and a lack of interference with subsequent viral cultures, and as part of studies that use chemical dyes to understand better the pathophysiology of ocular surface disorders, the staining characteristics and antiviral activity of sulforhodamine B and lissamine green B were investigated. METHODS: Staining of rabbit corneal epithelial cell cultures by sulforhodamine B and lissamine green B was compared to that of fluorescein and rose bengal. Diffusion of each dye through a collagen gel was measured. Uptake of lissamine green B by herpes simplex virus type 1 (HSV-1)-infected Vero cell cultures was compared at several times postinfection. The effect of sulforhodamine B and lissamine green B on HSV-1 plaque formation in Vero cells was determined. The cellular toxicity of sulforhodamine B and lissamine green B in vitro was examined by a quantitative 14C-amino acid uptake assay and by a qualitative cell viability assay. Finally, the effect of sulforhodamine B and lissamine green B on viral replication was compared in vivo with that of rose bengal in a rabbit model of herpetic epithelial keratitis. RESULTS: Rose bengal vividly stained cell monolayers of explant cultures of rabbit corneal epithelium. By light microscopy, sulforhodamine B and lissamine green B, like fluorescein, did not stain the epithelial cells, but did stain the corneal explant stroma. Pretreatment of epithelial cells with 0.25% trypsin for 5 minutes failed to induce dye uptake; however, pretreatment with 0.5% Triton X-100 for 5 minutes resulted in nuclear staining by lissamine green B, but not sulforhodamine B. When added to a collagen gel, the relative diffusion rate was fluorescein > lissamine green B > sulforhodamine B > rose bengal. By spectrophotometric analysis, HSV-1-infected and uninfected Vero cells bound equivalent amounts of lissamine green B until late in infection, when infected cells took up more dye (P < 0.001). A direct neutralization assay showed that 0.06% lissamine green B or 0.5% sulforhodamine B reduced HSV-1 plaque formation in Vero cells by greater than 50%, when present at the time of viral adsorption. By a quantitative 14C-amino acid uptake assay, lissamine green B was toxic to Vero cells in a dose-dependent manner, whereas sulforhodamine B was relatively nontoxic at the concentrations tested. By a cell viability assay, however, neither dye showed significant cellular toxicity. In a rabbit model of herpetic epithelial keratitis, rose bengal significantly reduced viral replication and recovery, whereas sulforhodamine B and lissamine green B had no effect. CONCLUSIONS: Neither sulforhodamine B nor lissamine green B stain healthy, normal cells. Lissamine green B stains membrane-damaged epithelial cells, but sulforhodamine B does not. Both sulforhodamine B and lissamine green B stain corneal stroma. Lissamine green B inhibits HSV-1 plaque formation at low concentrations of dye in vitro, which correlates with suppression of cellular metabolism as demonstrated by a 14C-amino acid uptake assay, but does not affect cell viability. Neither sulforhodamine B nor lissamine green B inhibit viral replication or recovery in vivo.

Bilateral electroretinographic changes induced by unilateral intra-visual cortex inoculation of herpes simplex virus type 1 in BALB/c mice.

Intra-visual cortex inoculation of 10(2) plaque-forming units of herpes simplex virus type 1 (KOS-63) induced physiologic and morphologic retinal changes in 62.3% (33/53) of infected animals; of these, 91% were bilateral. In contrast, inoculation of the same viral titers into the frontal lobe induced retinal alterations in only 13.3% (2/15). Initially, there was a decrease of the b-wave amplitude and retinal sensitivity and necrotic changes of the ganglion cells and nuclei in the inner nuclear layer. Immunoperoxidase staining for virus-specific antigens showed positive staining of the same cell type. Over time, there was a progressive decrease in the electroretinogram until it was extinguished and the retina was replaced by gliotic tissue. Parallel viral recovery studies demonstrated detectable infectious virus in one of eight eyes on day 2 after inoculation and in three of eight eyes on day 4. Thereafter, there was an increase in the percentage of eyes with infectious virus and a concomitant increase in viral titers. Immunoperoxidase staining of brain sections obtained on days 6 through 8 demonstrated virus-specific antigens on cells in the lateral geniculate nuclei and the suprachiasmatic nuclei bilaterally.

Herpes simplex virus type 1 infection of mouse astrocytes treated with basic fibroblast growth factor.

We explored a possible role for the basic fibroblast growth factor (FGF) receptor during herpes simplex virus type 1 (HSV-1) infection of primary mouse astrocytes, glial cells of the central nervous system known to express FGF receptors. Plaque reduction experiments showed that treatment of astrocyte monolayers with human recombinant basic FGF failed to inhibit HSV-1 infectivity, although treatment with either heparin or poly-L-lysine reduced it by approximately 100%. Identical results were obtained when monolayers of human embryonic lung fibroblasts or African green monkey kidney cells were treated with FGF, heparin or poly-L-lysine prior to HSV-1 infection. We conclude that the basic FGF receptor is not involved in the uptake of HSV-1 during productive infection of astrocytes. Our findings suggest that this molecule is not the predominant cellular receptor for HSV-1 among vertebrate cells and plays no role in defining HSV-1 neurotropism in vivo.

Direction of transneuronal transport of herpes simplex virus 1 in the primate motor system is strain-dependent.

We examined the axonal transport of two strains of herpes simplex virus 1 (HSV-1) within the central nervous system of cebus monkeys. Each strain was injected into the "arm area" of the primary motor cortex. One strain, HSV-1(McIntyre-B), was transported transneuronally in the retrograde direction. It infected neurons at sites known to project to the arm area of the primary motor cortex (e.g., ventrolateral thalamus). In addition, "second-order" neurons were labeled in the deep cerebellar nuclei (dentate and interpositus) and in the globus pallidus (internal segment). This result supports the concept that the arm area of the primary motor cortex is a target of both cerebellar and basal ganglia output. In contrast, the other strain, HSV-1(H129), was transported transneuronally in the anterograde direction. It infected neurons at sites known to receive input from the arm area of the primary motor cortex (e.g., putamen, pontine nuclei). In addition, "third-order" neurons were labeled in the cerebellar cortex (granule and Golgi cells) and in the globus pallidus (largely the external segment). Our observations suggest that strain differences have an important impact on the direction of transneuronal transport of HSV-1. Furthermore, it should be possible to examine the organization of cerebellar and basal ganglia loops with cerebral cortex by exploiting transneuronal transport of HSV-1 and virus strain differences.

Infection of human neural cell aggregate cultures with a clinical isolate of cytomegalovirus.

Human neural cell aggregate cultures were prepared from dissociated fetal brain tissue and maintained in rotation culture. After 35 days in culture, aggregates had the histologic appearance of dense, immature, neural cells in a tightly packed neuropil. Electron microscopy revealed ultrastructural features suggestive of immature neurons and neuroglia. In addition, neuron-specific enolase and glial fibrillary acidic protein associated with radial glial cells were detected within the aggregates by immunoperoxidase staining. When infected with a laboratory-adapted strain of cytomegalovirus (CMV), [AD169], cells containing large, bizarre, nuclei and CMV-induced intranuclear inclusion bodies were dispersed throughout the aggregates at 16 days postinfection. In situ hybridization using a CMV-specific DNA probe and electron microscopy confirmed the presence of virus sequences as well as virus particles at histologic sites of cytopathology. In sharp contrast, aggregate cultures infected with a CMV strain recovered from the retina of an acquired immune deficiency syndrome (AIDS) patient with CMV retinitis and encephalitis displayed distinct foci of cytopathology at 23 days postinfection, a pattern not observed in CMV [AD169]-infected aggregates. Our findings suggest that human neural cell aggregates represent a a promising multicellular non-neoplastic culture system in which to study the replication of human neurotropic viruses within neural tissue.

Induction of encephalitis in SJL mice by intranasal infection with herpes simplex virus type 1: a possible model of herpes simplex encephalitis in humans.

Herpes simplex encephalitis (HSE) is characterized by focal lesions of hemorrhage and necrosis, primarily in the inferior temporal lobe. Since immunosuppressed patients with HSE lack the focal inflammatory changes and temporal lobe localization, it has been suggested that the immune system participates in the pathogenesis of HSE. Evaluation of this hypothesis has been impeded by the lack of an immunologically defined animal model that resembles the human disease. Toward this end, 10 strains of inbred mice were infected intranasally with a neurovirulent clinical isolate of herpes simplex virus type 1. Most mice died without localizing signs of disease in the central nervous system. However, a significant number of SJL mice had a pattern of encephalitis highly reminiscent of that described in humans. To our knowledge, this is the first murine model that faithfully mimics this human disease, and thus it affords the opportunity to study the immunopathogenesis of HSE.