Spontaneous ocular shedding of HSV-1 in latently infected rabbits

The unscarified corneas of rabbits were inoculated with 50 microliter of 2-4 X 10(6) PFU/ml of herpes simplex virus, type 1 (HSV-1), McKrae strain in 10 separate experiments over a 12-month period. Sixty of 104 (57.7%) rabbits survived to postinoculation (PI) day 20. These sixty rabbits were swabbed with dacron-tipped swabs for twenty consecutive days (PI days 20-39). The tear film collected on the swabs was immediately placed in tissue culture tubes with confluent primary rabbit kidney (RK) cell monolayers. The RK monolayers were monitored for cytopathic effects indicative of HSV-1. Fifty-eight of the sixty rabbits (96.7%) inoculated had at least one positive episode. Ninety-three of the 120 eyes (77.5%) of the latently infected rabbits had at least one positive episode. Virus was detected in 72 of the 93 positive eyes (77.4%) between PI days 20 and 29 and in 21 of the 93 positive eyes (22.5%) between PI days 31-39. A total of 2400 swabs were taken and 324 were positive (13.5%). All of the 58 positive rabbits were used later for ocular induction of HSV-1 and all 116 eyes of the latently infected rabbits shed virus for at least four consecutive days during induction.     

Strain specificity of spontaneous and adrenergically induced HSV-1 ocular reactivation in latently infected rabbits

Spontaneous ocular shedding and adrenergic induction of ocular shedding were examined in rabbits infected with ten strains of herpes simplex virus type 1 (HSV-1): McKrae, KOS, F, Rodanus, 17 Syn+, RE, E-43, SC-16, MacIntyre, and CGA-3. All ocular inoculations were with 50 microliter of HSV-1 with titers between 1-10 X 10(6) PFU/ml. All corneas, except those that received the McKrae strain, were scarified. Acute ocular infection was determined by slit-lamp biomicroscopy. Dendritic keratitis or geographic ulcers developed in all eyes of all rabbits within 10 days after ocular inoculation. All eyes of all surviving rabbits were swabbed for 20 consecutive days during days 20-39 postinoculation (PI). On PI day 19, no active lesions were present as judged by slit-lamp biomicroscopy. Ocular tear film was collected on a Dacron-tipped swab and placed on primary rabbit kidney cell monolayers. The cell monolayers were monitored for cytopathic effects consistent with HSV-1 infection. Spontaneous HSV-1 shedding was detected in some eyes from all groups of latently infected rabbits, except those infected with CGA-3. Spontaneous shedding (positive swabs/total swabs) of the other nine strains ranged from 0.7% to 15.7%. After PI day 42, the rabbit eyes received 6-hydroxydopamine by iontophoresis, followed for 5 days by topical application of 2% epinephrine. This procedure results in induced HSV-1 ocular shedding for a duration of 3-5 days in rabbits infected with the McKrae strain. In rabbits latently infected with KOS, F, RE, MacIntyre, and CGA-3, no induced HSV-1 shedding was detected.(ABSTRACT TRUNCATED AT 250 WORDS)     

Quantitation and kinetics of induced HSV-1 ocular shedding

Iontophoresis of 6-hydroxydopamine (6-HD) to the rabbit eye, followed by topical instillation of 2% epinephrine, induces ocular shedding of herpes simplex virus type 1 (HSV-1) reliably and with a high frequency in latently infected rabbits. Rabbit eyes inoculated with HSV-1 (McKrae strain) showed dendritic lesions indicative of acute HSV infection and subsequently shed virus spontaneously at least once during days 20 to 39 postinoculation (P.I.). Two iontophoretic conditions were employed. Group A (3 rabbits, 60.3 days P.I.) received iontophoresis of 1.0% 6-HD at 0.75 mAmp for 3 min. Group B (three rabbits, 67.3 days P.I.) received iontophoresis of 0.1% 6-HD at 0.5 mAmp for 8 min. Following iontophoresis, 2% epinephrine was instilled topically once on the day of iontophoresis and twice daily for four consecutive days. Tear film was collected on Dacron swabs and titered on African green monkey kidney cells by a plaque assay procedure. In group A, 100% (6/6) of the eyes shed virus, and the average duration of shedding was 4.0 days. The titers ranged from 2.0 to 7.7 X 10(4) plaque-forming units (PFU) per eye. The highest daily average titer, 9.89 X 10(3) PFU/eye, occurred on day 5 following iontophoresis. In Group B, 100% (6/6) of the eyes also shed virus, and the average duration of shedding was 5.3 days. The viral titer of the tear film ranged from 5.0 to 1.4 X 10(5) PFU/eye. The highest daily average titer, 4.68 X 10(3) PFU/eye, also occurred on day 5 following iontophoresis.(ABSTRACT TRUNCATED AT 250 WORDS)     

HSV-1 retinitis and delayed hypersensitivity in DBA/2 and C57BL/6 mice

Following uniocular anterior chamber (a.c.) inoculation of HSV-1 into BALB/c mice, the architecture of the uninoculated eye is destroyed by a pan-retinal inflammatory process within 10 days. BALB/c inoculated with HSV-1 via the a.c. route also develop anterior chamber associated immune deviation (ACAID), one characteristic of which is impairment of virus-specific delayed hypersensitivity (DH) responsiveness. To determine whether other strains of mice develop ACAID and contralateral retinitis, DBA/2 and C57BL/6 mice were inoculated with HSV-1 (KOS strain) via the a.c. route. At intervals after inoculation, animals were examined clinically for evidence of retinitis in the uninoculated eye and were either (1) sacrificed for virus recovery studies or (2) ear-challenged for DH assays. Seven of 15 DBA/2 mice had clinical evidence of retinitis in the uninoculated eye by day 10 post-inoculation (p.i.). At this time, the average titer of virus in the uninoculated eyes was 3.37 log10 PFU/ml. Animals of this strain also had significantly impaired HSV-1 virus-specific DH responses consistent with ACAID. No (0/12) C57BL/6 developed contralateral retinitis; at day 10 p.i., the average virus titer in the uninoculated eyes of C57BL/6 mice was 1.10 log10 PFU/ml. Following a.c. inoculation of HSV-1, C57BL/6 mice developed vigorous DH responses comparable to those of subcutaneously immunized, DH-positive animals. The results of these experiments suggest that different strains of mice vary in their susceptibility to HSV-1 retinitis and that the development of retinitis is linked to the amount of virus in the uninoculated eye.(ABSTRACT TRUNCATED AT 250 WORDS)     

Induction of ocular herpes simplex virus shedding by iontophoresis of epinephrine into rabbit cornea

Ocular herpes simplex virus type 1 (HSV-1) shedding from the latently infected rabbit was induced by iontophoresis of 0.01% epinephrine into the eye. The iontophoresis of epinephrine was at 0.8 mAmp for 8 min once a day for 3 consecutive days. Shedding was determined by the presence of HSV-1 in the tear film obtained with eye swabs. Unilateral epinephrine iontophoresis performed 60 days after inoculation of the virus resulted in ipsilateral HSV-1 shedding in all cases (7/7). Bilateral epinephrine iontophoresis performed on selected days during 170 to 365 days after inoculation resulted in HSV-1 shedding in 75% of the eyes (21/28) and 100% of the rabbits (14/14). All shedding was initiated within 3 days after the third treatment with epinephrine iontophoresis. The shedding frequency induced by epinephrine iontophoresis was significantly higher (pb less than 0.05) than that induced by the other methods employed. HSV-1 was detected in one or both cocultivated explants of trigeminal and superior cervical ganglia for every eye in all experimental groups, indicating that all eyes had the potential to shed. In conclusion, epinephrine iontophoresis induced ocular HSV-1 shedding reliably and with a high frequency in the latently infected rabbits. Furthermore, we suggest that this easily reproducible model of viral shedding offers a system for studying the factors involved in recurrent HSV-1 ocular infections.     

Timolol induces HSV-1 ocular shedding in the latently infected rabbit

Timolol iontophoresis into the eye can induce herpes simplex virus type 1 (HSV-1) shedding in rabbits latently infected with HSV-1 strain McKrae. Anodal iontophoresis of 0.01% timolol was done at 0.8 mAmp for 8 min once a day for 3 consecutive days. Viral shedding was determined by the presence of HSV-1 in the preocular tear film obtained by eye swabs. In two experiments, iontophoresis of 0.01% timolol resulted in all eyes (18/18) shedding HSV-1 for an average duration of 4.3 days. When 5.0% timolol was applied topically to rabbit eyes supersensitized by iontophoresis of 6-hydroxydopamine (6-HD), all eyes (10/10) shed virus for an average duration of 2.9 days. All eyes (12/12) receiving iontophoresis of 6-HD, pre- and posttreatment with topical application of 5.0% timolol, and posttreatment with topical application of 1.0% epinephrine shed virus for an average duration of 3.6 days. Eyes treated with topical application of 5.0% timolol alone showed no difference in HSV-1 ocular shedding, compared with untreated eyes. We concluded that both iontophoresis of 0.01% timolol and topical application of 5.0% timolol to adrenergically supersensitized eyes induced HSV-1 shedding reliably and with a high frequency, and that topically applied timolol does not block the HSV-1 ocular shedding induced by epinephrine in adrenergically supersensitized eyes.     

Chorioretinal disease patterns in congenic mice following intraocular inoculation with HSV-1

Disease patterns and immunologic parameters were studied employing inbred and Igh-1 disparate congenic mice to determine the role of host genetics and Igh-1-linked gene products in the von Szily model of viral chorioretinitis. Following intracameral inoculation of 1.5 x 10(4) PFU HSV-1 (KOS), 100% of BALB/c (Igh-1a), 62% of A/J (Igh-1e) and none of the C57BL/6J (Igh-1b) inbred mice developed contralateral necrotizing chorioretinitis. Multigenic differences between inbred mice prohibit conclusions about the specific role of Igh-1-linked immune regulation in this model. In order to more exactly define Igh-1-specific restriction of HSV-1-mediated chorioretinitis, Igh-1-disparate, congenic BALB/c mice were studied following both anterior chamber and intravitreal inoculation protocols. Anterior chamber inoculation resulted in contralateral retinal necrosis in 75% of BALB/c (Igh-1a) mice, 30% of C.AL-20 (Igh-1d) and 5% of the C.B-17 (Igh-1b) congenic mice; all strains showed ipsilateral retinal sparing. Following intravitreal inoculation of HSV-1 a similar restricted disease pattern was found in contralateral eyes. Contralateral chorioretinitis developed in 30% of BALB/c, 15% of C.AL-20 and 6% of C.B-17 mice. Ipsilateral disease, however, was found in all murine strains. These disease patterns developed despite equivalent suppression of systemic DTH and equivalent RPE permissivity to viral replication. These data demonstrate that host genetics strongly regulates contralateral HSV-1-mediated chorioretinal disease patterns by a mechanism unrelated to the development of systemic suppression of DTH and specifically support a dominant role for gene products linked to the Igh-1 locus in the immunomodulation of ocular disease.     

Bilateral alterations of the ERG and retinal histology following unilateral HSV-1 inoculation

The physiological condition of the retinas of BALB/c mice inoculated unilaterally in the anterior chamber with the KOS strain of herpes simplex virus type 1 (HSV-1) was monitored by ERG recordings. After the ERG recordings, the retinas were examined for histopathological changes. In the inoculated eye, depressed ERGs were recorded on day 2 PI and abolished ERGs on day 4 PI. The changes in the ERGs were complete by day 5-6 PI. Of the 53 inoculated eyes followed for longer than day 6 PI, four (7.5%) remained normal, 30 (56.6%) had reduced ERGs and 19 (35.8%) had abolished ERGs. In the contralateral eyes, the first changes were noted on day 8 PI, and abolished ERGs were recorded on day 9 PI. Of the 55 contralateral eyes followed for longer than 10 days, 15 (27.3%) remained normal, four (7.2%) had reduced ERGs and 36 (65.4%) had abolished ERGs. The percentage of eyes with depressed ERGs was significantly higher in the inoculated than in the uninoculated eyes, and the percentage of eyes with abolished ERGs was significantly higher in the uninoculated eyes than in the inoculated eyes. The histopathological alterations were different for the two eyes. In the inoculated eyes, the changes were mainly in the outer retina, with characteristic folds in the photoreceptor and outer nuclear layer interspersed with normal appearing retina. The pigment epithelium was also abnormal. In the uninoculated eyes, the changes began in the inner retina but rapidly spread to all layers of the retina. This panretinal necrosis accounted for the higher percentage of abolished ERGs in the uninoculated eyes. The differences in the alterations of the ERG and the histopathological changes may be related to the underlying mechanism of action of the HSV-1 during the evolution of the experimental retinopathy.     

Subconjunctival antisense oligonucleotides targeting TNF-α influence immunopathology and viral replication in murine HSV-1 retinitis

BACKGROUND: To investigate the role of tumor necrosis factor-alpha (TNF-alpha) in immunopathology and viral replication in the contralateral eye in the von Szily model of herpes simplex virus (HSV)-1 acute retinitis. METHODS: In vivo distribution was analyzed after subconjunctival injection of FITC-labeled antisense oligonucleotides (ASON). After HSV-1 (KOS) was injected in the right anterior chamber (AC) in BALB/c mice, the course of the contralateral retinitis was evaluated. The left eyes were treated with either TNF-alpha ASON, sequence-unspecific control (CON), or buffer. The ocular TNF-alpha content was quantified by ELISA. The delayed-type hypersensitivity (DTH) reaction, uptake of [3H]thymidine from regional lymph nodes (rln)- and spleen cells, serum-neutralizing antibodies, and viral titer in the eyes were evaluated. RESULTS: After subconjunctival injection, FITC-labeled ASON were found in the choroid and retina. In the TNF-alpha ASON-treated eyes, TNF-alpha expression and the incidence and severity of retinitis were reduced on day 8 postinfection (PI) (p < 0.05). On day 10 PI, higher viral titers were only seen in the eyes of the TNF-alpha ASON group (p < 0.05), and retinitis was slightly more severe on day 12 PI. While the HSV-1 specific [3H]thymidine uptake from rln cells was higher in the TNF-alpha ASON mice (p < 0.05), the [3H]thymidine uptake from spleen cells, the DTH response, and the neutralizing-antibody titers did not differ between the groups. CONCLUSIONS: After regional blockade of TNF-alpha in experimental HSV-1 retinitis TNF-alpha seems to possess an antiviral capacity against HSV-1 in the contralateral eye and participates in the immunopathology of HSV-1-induced acute retinitis.     

Effects of serotonergic compounds on aqueous humor dynamics in monkeys

PURPOSE: The effects of several serotonergic agonists on aqueous humor formation (AHF), total outflow facility (OF) and intraocular pressure (IOP) were investigated in living cynomolgus monkeys. METHODS: We determined the effect of a single topical unilateral 300 microg or 3 mg dose of the 5-HT agonists serotonin, 5-carboxamidotryptamine (5-CT), sumatripan, gepirone, and 8-hydroxy-2(di-n-propylaminotetralin) (8-OH-DPAT) and a 450 microg dose of flesinoxan on IOP (Goldmann applanation tonometry), AHF (scanning ocular fluorophotometry) and total OF (8-OH-DPAT only, topically and intracamerally). RESULTS: Serotonin, 5-CT, sumatripan or gepirone had no significant effect on IOP or AHF. 8-OH-DPAT caused an AHF increase of approximately 70% over 6 hr in both ipsilateral drug- and contralateral vehicle-treated eyes, but no significant change in IOP compared with baseline measured on a separate occasion in the same animals. 8-OH-DPAT did not increase protein levels or rate of entry of systemically administered fluorescein in the anterior chamber aqueous humor compared to historic controls, and no difference was seen between ipsilateral and contralateral eyes. Flesinoxan had no effect on IOP and produced an insignificant 25% increase in flow in treated eyes compared to baseline. CONCLUSION: The results for 8-OH-DPAT and possibly flexinoxan indicate the presence of a secretion-stimulating 5-HT1A receptor in monkey ciliary epithelium that has little effect on IOP. OF was unchanged following 8-OH-DPAT administered topically or following intracameral exchange.     

Fuchs heterochromic cyclitis: rubella virus antibodies and genome in aqueous humor

PURPOSE: To characterize the polyspecific intraocular antibody synthesis in aqueous humor of patients with chronic inflammatory diseases of the eye and to detect the causative antigen in Fuchs heterochromic cyclitis (FHC). DESIGN: Retrospective case-control study. METHODS: Intraocular antibody synthesis is detected in aqueous humor with the Antibody Index [AI] (improved Goldmann-Witmer Index) and quantified as specific antibody fraction, F(s) (intraocular specific antibody concentration in percent of intraocular total immunoglobulin G in aqueous humor). Virus detection is by nested polymerase chain reaction. RESULTS: Fifty-two eyes of 52 patients with clinically defined FHC (aged 16-73 years) had an intraocular synthesis of rubella antibodies (AI > or =1.5). The rubella genome was detected in 5 (18%) of 28 aqueous humor samples investigated, or in 5 (56%) of 9 patients aged <40 years. Oligoclonal IgG was synthesized in 34 (87%) of 39 eyes. Unaffected fellow eyes (n = 3) or cerebrospinal fluid (n = 2) were normal. In FHC the median rubella AI = 20.6 (total range 1.5-309) was seven times higher than in multiple sclerosis (MS) patients (n = 15) with uveitis intermedia or periphlebitis retinae. In MS the intraocular rubella antibody synthesis (frequency 73%) is part of a polyspecific immune response (increased measles AI in 80%, varicella zoster virus AI in 47%, herpes simplex virus AI in 23%). The median rubella-F(s) = 2.6% in FHC (range = 0.14%-45.9%) was approximately 40 times higher than in MS, consistent with a virus-driven antibody response in FHC. Noninflammatory controls (50 senile cataracts) had neither an intraocular rubella antibody synthesis (normal AI < or =1.4) nor rubella antigen in aqueous humor. The rubella AI was normal in all patients with an intraocular toxoplasmosis (n = 24), anterior uveitis (n = 27), herpes simplex virus iritis (n = 25), and varicella zoster virus iritis (n = 14). CONCLUSIONS: Fuchs heterochromic cyclitis is a rubella virus-driven disease with persistence of the virus preferentially detected in the younger patients. The proposed laboratory supported diagnosis of FHC is based on the increased rubella Antibody Index. The virus etiology gives a rationale for omitting the ineffective corticosteroid therapy of FHC.     

Insulin and tropicamide accumulate in the contralateral, untreated eye of rats following ipsilateral topical administration by a mechanism that does not involve systemic uptake.

BACKGROUND: This study was designed to determine: (1) whether the accumulation of insulin in the contralateral retina and aqueous humor following ipsilateral topical insulin administration was due to systemic uptake and (2) whether tropicamide, applied to one eye, could induce dilation in the contralateral eye by a mechanism that did not involve systemic uptake. METHODS: Insulin eye drops were applied to the left eye of intact and decapitated rats, and their retinas and aqueous humors were then removed and their insulin levels quantified. In a separate experiment live animals received 0.1% tropicamide in their left eye and had their pupillary dilation response in both eyes measured at different time points. RESULTS: Administration of insulin to the left eye of decapitated rats resulted in its significant accumulation not only in the left retina and aqueous humor, but also in the retina and aqueous humor of the right eye. Similar aqueous humor results were obtained when live animals were used. Tropicamide drops induced marked pupillary dilation in treated eyes; the pupils of the contralateral, untreated eyes also dilated significantly, but less than did the treated pupils. The pupils of rats injected with tropicamide intravenously showed negligible dilation. CONCLUSIONS: These results showed that insulin accumulated in the retina and aqueous humor of contralateral, untreated eyes following topical application, by a mechanism that did not appear to involve systemic uptake. Similarly, tropicamide provoked a dilation response in the unheated eye by a mechanism that similarly did not appear to involve uptake from the blood.     

An immunogenetic analysis of resistance to herpes simplex virus retinitis in inbred strains of mice

Specific inbred strains of mice have been shown to vary considerably in their resistance and susceptibility to herpes simplex virus (HSV) infection. We injected 2 X 10(5) plaque forming units (PFU) of the KOS strain of HSV-1 intracamerally into one eye of BALB/c, C57Bl/6, and F1 (BALB/c X C57Bl/6) mice. HSV-1 antigens were localized in frozen sections of enucleated eyes at 10 to 14 days post-inoculation. Injected eyes of BALB/c mice showed an anterior uveitis with HSV-1 antigens in the anterior segment and an intact retina free of HSV antigens. The retina of the contralateral uninjected eye was necrotic and contained HSV-1 antigens. In both C57Bl/6 and F1 mice, HSV antigens were limited to anterior segment structures in the injected eye, whereas, in contrast to BALB/c mice, the contralateral retina appeared histologically normal and contained no viral antigens. The C57Bl/6 and F1 strains remained relatively resistant to retinal infection even if pretreated with up to 800 Rads of irradiation. The retinas of normal or sublethally irradiated C57Bl/6 and F1, but not BALB/c strains, were also resistant to intravitreal injection of HSV. These results suggest that resistance to HSV retinitis is a dominantly inherited trait, which depends only partly upon immunologic factors and may be heavily influenced by the inherent ability of host cells from different murine strains to support a productive viral infection.     

The effect of bunazosin hydrochloride on intraocular pressure and aqueous humor dynamics in human]

The effects of bunazosin hydrochloride, a new highly selective alpha 1 adrenergic antagonist, on the intraocular pressure (IOP) and aqueous humor dynamics in healthy human volunteers were studied. Unilateral topical administration of 0.1% bunazosin significantly lowered IOP from 1 to 10 hrs in bunazosin-treated eyes, and from 2 to 8 hrs in contralateral placebo-treated eyes, with its maximum reduction at approximately 3 hrs, when the IOP decreased 5.0 mmHg in treated eyes, and 3.5 mmHg in contralateral eyes from the baseline, respectively. Fluorophotometrically measured aqueous humor flow rates did not change significantly either in treated or contralateral eyes. Bunazosin caused significant reduction of systolic and diastolic blood pressure 5 hr after application. The difference in pupil diameter between treated and contralateral eyes was found to be statistically significant from 1 to 5 hrs. A small but significant increase in anterior chamber depth also was observed from 2 to 6 hrs. Pulse rate and refraction were not substantially altered. There was moderate conjunctival vessel dilation, however, no serious complications were encountered. The results suggest that bunazosin hydrochloride appears to have great clinical potential for ocular hypotensive therapy.     

Apolipoprotein E modulates establishment of HSV-1 latency and survival in a mouse ocular model

PURPOSE: To evaluate and compare the neuroinvasiveness and neurovirulence after ocular HSV-1 infection in ApoE knockout (ApoE-/-) and control C57BL/6 (ApoE+/+) mice. METHODS: Age-matched (14 weeks of age) C57BL/6J (ApoE+/+) female mice and female ApoE knockout (ApoE-/-) mice were inoculated by corneal scarification with HSV-1 strain 17Syn+. Analysis of HSV-1 replication in the mouse cornea was assessed through infectious virus assays of ocular (tear film) swabs at 1 to 5 days postinoculation (PI), slit-lamp examination (SLE) of corneas at PI days 1 to 7, and survival of infected mice. The contribution of apoE to the efficient establishment of latency was measured by real-time PCR quantitation of the latent viral genome in the trigeminal ganglia (TG) of infected mice. RESULTS: These studies showed that HSV-1 strain 17Syn+ replicates efficiently in the eyes, regardless of the host ApoE genotype. Neither the scoring of corneal pathology via SLE nor the infectious virus assay of the tear film resulted in any statistical differences between ApoE knockout (-/-) mice or the C57BL/6 (ApoE+/+) mice. In mice latently infected with HSV-1, our real-time PCR data showed significantly lower viral copy numbers of HSV-1 DNA in ApoE knockout (ApoE-/-) mice compared with C57BL/6 (ApoE+/+) mice. C57BL/6 (ApoE+/+) mice are more susceptible to the neurovirulence of HSV-1 strain 17Syn+ than female ApoE knockout (-/-) mice, as demonstrated by the fact that 50% (7/14) of the female C57BL/6 (ApoE+/+) mice inoculated with 17Syn+ died, as opposed to none (0/14) of the age- and sex-matched ApoE knockout mice. CONCLUSIONS: These data indicate that age (14 weeks) and sex-matched (female) wild mice with an ApoE null background (ApoE-/-) are more resistant and less efficient in the establishment of latency compared with ApoE+/+ mice in the C57BL/6 background.     

Effect of aminoguanidine, a nitric oxide synthase inhibitor, on ocular infection with herpes simplex virus in Balb/c mice

PURPOSE. To study the effect of aminoguanidine (AMG), an inhibitor of nitric oxide production, on the ocular infection of Balb/c mice with herpes simplex virus (HSV) type 1 strain F and HSV-2 strain G. METHODS. Animals were treated with different amounts of AMG (0.5, 0.1, and 0.05 mg/mouse) by topical application in the eye from postinfection (PI) days -2 through +5, considering 0 the day of infection. At different PI days, development of herpetic keratitis was evaluated in treated and control mice. RESULTS. Treated animals showed a dose-dependent increase in ocular disease after viral infection, compared with control animals. Viral titers in ocular washings were higher in AMG-treated mice (PI day 2, HSV-1: AMG 0.5 mg, 1.3 x 10(3) plaque-forming units (PFU)/ml; control, 0. 22 x 10(2) PFU/ml, P < 0.025). At PI day 3, control corneas had only scattered inflammatory cells, whereas those from treated animals showed a conspicuous infiltrate consisting primarily of neutrophils. Viral titers were also higher in brains of treated mice. These animals died earlier and in a greater proportion than control animals (percentage of mortality, PI day 12, HSV-1: AMG 0.5 mg, 40% +/- 4%; control, 18% +/- 3%, P < 0.05). CONCLUSIONS. These data indicate an inhibitory effect of nitric oxide on HSV ocular infection.     

Further studies of the ipsilateral and contralateral responses to topical nitrogen mustard.

The short term ipsilateral and contralateral responses of the rabbit eye to topical 1% nitrogen mustard were studied by measurement of protein in the aqueous humor, anterior segment fluorescein angiography, and aqueous fluorophotometry. The ipsilateral response was characterized by intense miosis, ischemia of the iris, and leakage of fluorescein (and presumably protein) into the posterior chamber. The iris ischemia was at least partly secondary to the pronounced elevation of intraocular pressure that occurs following nitrogen mustard. A contralateral rise in protein levels in the aqueous humor was seen in five of 22 rabbits. No contralateral iris ischemia or miosis was observed. Neither the ipsilateral nor the contralateral response was inhibited by pretreatment with aspirin. Aqueous fluorophotometry was only slightly more sensitive in detecting a contralateral response than simple measurement of protein levels in the aqueous.     

Trifluridine decreases ocular HSV-1 recovery, but not herpetic lesions after timolol iontophoresis

To determine the effect of a topically applied antiviral agent on shedding of herpes simplex virus type 1 (HSV-1) into the tear film and corneal epithelial lesions, ten rabbits latently infected with HSV-1 were subjected to transcorneal iontophoresis of 0.01% timolol once a day for 3 consecutive days to induce viral shedding and lesions. Iontophoretic induction was performed similarly in five uninfected rabbits as controls. Half of the infected rabbits and all of the uninfected controls received topical 1.0% trifluridine five time a day for 9 days, beginning the day after the first iontophoresis. All eyes were examined daily for 10 days by slit-lamp biomicroscopy and tear film samples collected on swabs were analyzed for virus. In the infected rabbits, the eyes treated with trifluridine had significantly fewer swabs positive for HSV-1 than the untreated eyes (P less than 0.001); however, there was no significant difference in the numbers of lesions in the treated and untreated eyes. The uninfected controls had no positive swabs and developed no lesions. These results suggest that topical treatment with trifluridine may reduce recovery of HSV-1 from the tear film, but does not affect the incidence of iontophoretically induced corneal epithelial lesions.     

房水病毒载量和细胞因子检测在急性视网膜坏死诊断与治疗中应用的临床研究

目的探讨房水病毒载量和细胞因子检测在急性视网膜坏死(ARN)诊断治疗中的作用。 方法回顾性分析2017年3月至2021年7月于北京地坛医院眼科就诊ARN患者10例(13只眼)的病例资料。其中，男性8例(10只眼)，女性2例(3只眼)；年龄24~66岁，平均年龄(35.1±10.2)岁。采集所有患者的房水，明确病毒种类。将患者治疗前房水中病毒载量高于10⁵拷贝/ml定义为高载量组，低于者定义为低载量组。对所有患者均行静脉和玻璃体腔注射更昔洛韦治疗；对视网膜脱离者行玻璃体切除视网膜复位术或视网膜激光光凝术。检查所有患者治疗前后的最佳矫正视力、眼前节及眼底情况。采用聚合酶链式反应检测所有患者治疗前和每次随访时房水的病毒载量；采用流式细胞微球捕获芯片技术检测患者治疗前和随访时房水白细胞介素(IL)-6及IL-8的含量。房水病毒载量的下降速度以log(病毒载量)拷贝/ml表示。最佳矫正视力以例数和百分比表示；注射次数、房水病毒载量、IL-6及IL-8以中位数和四分位数表示。 结果ARN患者10例(13只眼)均为感染水痘带状疱疹病毒(VZV)导致，治疗后视网膜病变均得到有效控制，玻璃体腔注射次数为2~9次，平均中位数和四分位数6(5，9)次。继发视网膜脱离者4例(5只眼)，占38.46%(5/13)。其中，行玻璃体切除术者3例(3只眼)，占60.00%(3/5)；行视网膜激光光凝术1例(2只眼)，占40.00%(2/5)。患者治疗前房水VZV平均载量的中位数和四分位数为2.69×10⁵(4.27×10⁴，9.26×10⁷)拷贝/ml；治疗2周后病毒载量开始下降，logVZV平均每周下降(0.67±0.30)拷贝/ml；治疗6周后升高者3例(5只眼)，占38.46%(5/13)。患者治疗前房水IL-6和IL-8平均含量的中位数和四分位数分别为2993.3(1655.9，18751.3)pg/ml和253.6(195.8，1682.2)pg/ml；治疗2周后分别为398.1(251.2，2511.8)pg/ml和63.1(15.8，125.9)pg/ml，较治疗前明显下降。高载量组和低载量组患者的病程分别为(14.00±4.47)d和(11.33±8.26)d，前者时间长于后者19.07%。高载量组和低载量组患者平均注射次数的中位数和四分位数分别为7(6，9)和6.5(4，9)。高载量组和低载量组患者终末最佳矫正视力≥0.1者分别为2例(2只眼)和4例(4只眼)，分别占15.38%(2/13)和30.77%(4/13)；<0.1者分别为4例(5只眼)和2例(2只眼)，分别占38.46%(5/13)和15.38%(2/13)。高载量组和低载量组继发视网膜脱离者分别为4例(4只眼)和1例(1只眼)，分别占30.77%(4/13)和7.69%(1/13)。 结论ARN患眼房水的病毒载量在治疗后，经历平台期和对数下降期，并最终低至可检测阈值以下。房水IL-6和及IL-8含量可以评估眼内炎的严重程度。房水检测可以明确病毒种类，监测炎症反应和治疗效果，对于ARN的诊断与治疗具有重要参考价值。     

Corneal nerves contain intra-axonal HSV-1 after virus reactivation by epinephrine iontophoresis

Experimental ocular models of herpes simplex virus type 1 (HSV-1) reactivation have been used to monitor viral shedding in the tear film and the appearance of corneal epithelial lesions, but the temporal correlation between reactivation and the presence of viral particles in the corneal nerves has not been made. Two New Zealand white rabbits were inoculated with 20 microliters of HSV-1 McKrae strain (5.0 x 10(6) PFU/ml) in each eye. Beginning on postinfection day 82, ocular iontophoresis (0.8 mAmps for 8 min) of 0.01% epinephrine was done once a day for 3 consecutive days to induce reactivation. Ten limbal nerves from four corneas processed for transmission electron microscopy contained 883 unmyelinated and 40 myelinated axons. Seven nerves were positive for virus. Viral particles were found only in unmyelinated axons, and in low frequency (24/883). Virus was not found in Schwann cells, perineurium, or adjacent stroma nor were virus particles seen exiting axons. No enveloped virions were found. Axons from six nerves of four control corneas from rabbits with latent, but not reactivated, HSV-1 did not contain virus particles. Induction by corneal iontophoresis of epinephrine suggests that HSV-1 is translocated from the ganglion to the cornea through axonal transport mechanisms. For the first time, evidence of anterograde, intra-axonal transport of HSV-1 particles in response to epinephrine reactivation is demonstrated.