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PubMed1991

Hepatic drug metabolism during development in food-restricted female Sprague-Dawley rats.

Hepatic drug metabolism was investigated in female Sprague-Dawley rats fed ad libitum (A) or a restricted diet (R) (implemented from age 1 month), at 1.5, 4.5 and 12 months to determine the short- and long-term effects of caloric restriction. Microsomal cytochrome P-450 content and NADPH cytochrome c reductase activity were not modified by age. While dietary restriction did not affect cytochrome P-450, it significantly increased NADPH cytochrome c reductase activity at all time periods when compared to corresponding A-fed groups. Aniline hydroxylase and aminopyrine N-demethylase activity tended to decrease with age in the A-fed groups but the differences did not prove to be statistically significant. A significant decrease of aminopyrine N-demethylase was observed with age in R rats. A significant reduction of aniline hydroxylase activity was noted in the R groups compared to age-matched A-fed controls. In contrast, aminopyrine N-demethylase activity increased significantly, but only at 1.5 months of age. Glutathione S-transferase activity was augmented between 1.5 and 4.5 months of age, and this was followed by a significant decrease at age 12 months in both A and R groups. Dietary restriction had no effect on this enzymatic activity. The microsomal cholesterol and phospholipid content as well as the cholesterol/phospholipid molar ratio changed significantly between 1.5 and 4.5 months of age but not between 4.5 and 12 months of age. These parameters were unaltered by dietary restriction. In conclusion, in the female Sprague-Dawley rat there are no statistically significant changes in hepatic microsomal components and drug metabolizing capacity between 1.5 and 12 months of age. Dietary restriction resulted in significant changes in enzymes related to drug metabolism which varied with the enzyme examined. In general, these changes were similar after short- or long-term dietary intervention.

Aging↗

PubMed1990

Effects of energy restriction on norepinephrine turnover and serum glucose and fatty acids in lean mice.

Norepinephrine (NE) turnover rate was determined in several tissues of 5-week-old female mice fed a high carbohydrate diet (58% of energy as carbohydrate, 30% fat) either ad lib or restricted to 34 or 24 kJ/day (36 to 50% restriction) presented as 1 or 2 daily meals. When the restricted intakes were divided into 2 equal meals, daily NE turnover did not differ from that of ad lib-fed mice. When the above restricted amounts were provided as a single daily meal at the beginning of the dark period, NE turnover was 38% and 46% lower, respectively, in the heart only compared to ad lib-fed controls. Serum glucose and total free fatty acids were affected by dietary conditions known to produce sympathetic activation (high carbohydrate and high fat diets) and suppression (high protein diet and energy restriction as a single meal), but the changes were unrelated to fractional NE turnover. Thus, the lower NE turnover seen when food intake is restricted is due to the prolonged overnight fast and not due to the lower energy intake per se, and is not associated with serum concentration of glucose or total free fatty acids.

Animals↗

PubMed1976

Phenotypic mixing between N- and B-tropic murine leukemia viruses: infectious particles with dual sensitivity to Fv-1 restriction.

In effort to understand how N or B tropism is determined in murine leukemia virus (MuLV) particles, we analyzed the MuLV produced after dual infection of mouse cells by N- and B-tropic MuLV. The progeny MuLV from such a mixed infection are sensitive to Fv-1 restriction in both N- and B-type cells, but are still highly infectious for mouse cells which do not exhibit Fv-1 restriction. This dual sensitivity to Fv-1 restriction is a phenotypic property of MuLV produced by mixedly infected cells, since individual virus clones derived from this MuLV are either N- or B-tropic. In further experiments, we superinfected murine sarcoma virus (MSV)-transformed cells with mixtures of N- and B-tropic MuLVs. The rescued MSV is restricted in its ability to transforms both N- and B-type cells. The results suggest that N- and B-tropic MuLVs specify different determinants, which are incorporated into virions along with the viral genome and which are the recognition sites for Fv-1 restriction. The presence of a given determinant in a virion renders the virus sensitive to restriction in cells of the opposite Fv-1 type.

Cell Line↗

PubMed1977

Loss of Fv-1 restriction in Balb/3T3 cells following infection with a single N tropic murine leukemia virus particle.

The ability of various murine leukemia viruses (MuLVs) to replicate in mouse cells exhibiting Fv-1 restriction was analyzed by quantitative dose-response assays. In particular, the effect of infection with N, B, or NB tropic MuLVs on Fv-1b restriction in Balb/3T3 cells was measured with an infection center technique in which pseudotypes of murine sarcoma virus (MSV), which have been shown to exhibit Fv-1 dependence of expression, were used to quantitate the degree of restriction. The resulting dose-response curves indicate that productive infection of a single Balb/3T3 cell with N tropic MSV requires co-infection with two MuLV particles. These two MuLV particles are functionally distinguishable. One of them must be N tropic and must be added less than 18 hr after infection with N tropic MSV. The second MuLV particle, on the other hand, need not be N tropic and may be added at any time. Balb/3T3 cultures infected with sufficient N tropic MuLV become fully permissive to transformation by N tropic MSV and to productive infection by N tropic MuLV. This effect, termed "abrogation" of Fv-1 restriction, results from infection of a Balb/3T3 cell with a single N tropic MuLV particle, but apparently occurs without viral replication. It seems probable that a requirement for abrogation of Fv-1b restriction by a single infectious particle of N tropic MuLV, which does not itself replicate, is responsible for the two-hit dose-response relationship observed in infectivity titrations of N tropic MuLV in Balb/3T3 cells. The requirements that N tropic MuLV be added within a specified time period with regard to N tropic MSV in order for abrogation to occur suggests that in the absence of N tropic MuLV, the cellular Fv-1b restriction mechanism inactivates N tropic MSV by 9 hr after infection.

Cell Line↗

PubMed1993

HLA-B27-restricted CD8 T cells derived from synovial fluids of patients with reactive arthritis and ankylosing spondylitis.

Ankylosing spondylitis and seronegative spondylarthropathies such as Reiter's syndrome and reactive arthritis are strongly associated with HLA-B27. However, the mechanisms by which HLA-B27 is involved in disease susceptibility and pathogenesis are unknown. If the disease association is a consequence of HLA-B27's physiological function in antigen presentation, the disease should be mediated by cytotoxic T lymphocytes (CTLs) that recognise bacterial or self peptides presented by HLA-B27. Proof of this arthritogenic peptide model requires isolation of B27-restricted CD8 T cells from arthritic joints of patients with spondylarthropathies. An important question is whether "arthritogenic" bacteria such as yersinia or salmonella can generate HLA-B27-restricted bacteria-specific CTLs. We describe such HLA-B27-restricted CTLs. We tested a panel of 354 alpha beta-TCR CD8 T lymphocyte clones (TLCs) that had been derived from the synovial fluid of 4 patients with reactive arthritis and 2 patients with ankylosing spondylitis. In 1 patient with yersinia-induced arthritis, 2 TLCs were identified that killed specifically yersinia-infected B27 target cells. In another patient with salmonella-induced arthritis, 1 B27-restricted CD8 TLC that recognised both salmonella and yersinia was identified. In 5 of the 6 patients autoreactive CTLs were found, 5 of which showed B27-restricted killing of uninfected cell lines. B27-restricted CTLs with specificity for arthritogenic bacteria or autoantigens provide a missing link in the pathogenesis of the HLA-B27-associated spondylarthropathies.

Adult↗

PubMed1995

Intramolecular immunodominance and intermolecular selection of H2d-restricted peptides define the same immunodominant region of the measles virus fusion protein.

The generation of a sustained antibody response requires the participation of MHC class II-restricted T helper cells. We have identified class II-restricted sequences by immunizing BALB/c (H-2d) mice with 108 overlapping synthetic pentadecapeptides covering the whole sequence of the measles virus fusion protein (MV-F). Several strong T cell epitopes were found including a major cluster of H-2d-restricted peptides between amino acids 256 and 305. Some of these peptides including peptide F(421-435) and F(256-270) induced MV-specific T lymphocytes in vivo while other H2d-restricted MV-F sequences did not. Immunization with mixtures of selected peptides indicated a hierarchy among H2d-restricted sequences due to competition between peptides. The dominant peptide F(421-435) impaired the response to other T cell epitopes including F(256-270). The response to F(91-105) was obliterated by F(421-435) and F(256-270) but not by peptides devoid of a T cell epitope. When BALB/c mice were immunized with the MV, the immunodominant sequence F(421-445) was identified which included the synthetic peptide F(421-435). Our data suggest that competition during processing and/or presentation between H2d-restricted peptides defines the immunodominant sequence of the viral protein. Even though only a single immunodominant region was defined after immunization with the MV, peptides from other regions were able to induce MV-specific T cell responses. This finding is of interest for the design of subunit vaccines in general and for studying MV-specific T helper cells in an animal model in particular.

Amino Acid Sequence↗

PubMed1994

Effects of dietary restriction on induction of unscheduled DNA synthesis (UDS) and replicative DNA synthesis (RDS) in rat liver.

The effects of dietary restriction on the induction of unscheduled DNA synthesis (UDS) and replicative DNA synthesis (RDS) were studied in the hepatocytes of F344 rats exposed in vivo to dimethylnitrosamine (DMN) or CCl4. The animals were given food ad libitum, a restricted amount of food (4 g/rat/overnight) or no food. Hepatocytes were isolated 2 h after oral administration of DMN at a dose of 5 mg/kg body weight and 48 h after oral administration of CCl4 at a dose of 400 mg/kg body weight, and incubated for 4 h in Williams' medium E supplemented with either [3H]thymidine for UDS or 5-bromodeoxyuridine for RDS. UDS was determined by autoradiography and RDS was determined by the immunoenzymatic staining method. The background levels of UDS (net grains/nucleus) and RDS (cells in S phase) in control were -12.4 and 0.64% for ad libitum feeding, -6.8 and 0.04% for restricted feeding, and -8.1 and 0% for fasting. UDS induced by DMN and RDS induced by CCl4 were 19.4 and 3.3% for ad libitum feeding, 34.5 and 10.4% for restricted feeding, and 47.8 and 15.1% for fasting. DMN demethylase activity in rat liver was also found to increase with dietary restriction. These results indicate that dietary restriction modulates the responses of UDS and RDS in the liver of rats.

Animals↗

PubMed1991

Multiple sclerosis patients have reduced HLA class II-restricted cytotoxic responses specific for both measles and herpes virus.

It has been previously demonstrated that the generation of measles virus (MV)-specific cytotoxicity (CTL) is reduced in patients with multiple sclerosis (MS). By contrast, CTL specific for influenza virus (FLU) and mumps virus is normal. It is uncertain if reduced CTL is limited to MV in MS patients, or if reduced CTL may be found to other viruses as well. Since MV-specific CTL is predominantly restricted by HLA class II molecules, while FLU-specific and mumps-specific CTL have large HLA class I-restricted components, reduced MV-specific CTL may reflect a broader reduction in HLA class II-restricted CTL in patients with MS. To examine this question we studied the generation of CTL specific for herpes simplex virus type I (HSV). HSV-specific CTL, like MV-specific CTL is predominantly restricted by HLA class II molecules. We found that patients with MS had reduced generation of CTL to both MV and HSV. Most, but not all patients who had reduced generation of CTL to one virus also had a similar impairment with respect to the second virus. Some patients, however, had a reduction in the generation of CTL only to MV or to HSV. These findings extend our earlier observations regarding reduced MV-specific CTL in patients with MS to a second HLA class II-restricted virus, HSV. Such a reduction may reflect discrete impairments in immune function to separate viruses, possibly those that are associated with viral persistence, or may reflect a more generalized defect in HLA class II-restricted CTL.

Adult↗

PubMed1985

Homozygous typing cell-defined HLA-Dw specificities correlate better than serologically defined HLA-DR specificities with restriction elements for influenza virus-specific proliferative human T lymphocyte clones.

Human T lymphocyte clones with specific proliferative response to influenza A virus were derived by limiting dilution from peripheral blood lymphocytes (PBL) after in vitro stimulation with autologous irradiated, virus-infected PBL. Four OKT3+4+8- T lymphocyte clones (TLC) that showed HLA-restricted antigen-specific proliferative responses were used for a detailed analysis of the restriction elements for antigen presentation. None of the clones showed alloreactivity and all required the presence on the antigen-presenting cell of HLA class II antigens of one or other haplotype of the donor. Restriction elements for two clones were correlated with Dw1 rather than DR1, and for two others with Dw6 rather than DRw6. These latter clones showed differential recognition of HLA-Dw6 subtypes as defined tentatively by homozygous typing cells, without relationship to putative serological "splits" of DRw6. One of the Dw6-restricted clones was specific for a Dw6.1 (now Dw18) "subtype," confirmed by family segregation analysis, the other for a broad Dw6 (Dw18 and Dw19) specificity. Studies with a panel of monoclonal antibodies against monomorphic determinants of HLA class II antigens revealed heterogeneous patterns of blocking activity, distinguishing between clones of different restriction specificity. Inhibition patterns were partly as predictable from the known activity of the monoclonal antibody in alloantigeneic PLT systems. These results provide evidence that certain structures that function as restriction elements for antigen presentation also carry alloantigeneic determinants.

Antibodies, Monoclonal↗

PubMed1987

Importance of HLA-DQ and -DP restriction elements in T-cell responses to soluble antigens: mutational analysis.

We describe a new approach to delineating the restriction elements used by antigen-specific human T-cell lines. EBV-transformed B cell lines and congenic HLA class II antigen-loss mutants are used to present soluble antigen to immune T cells. In this way it is possible to assess the independent contribution of individual class II loci to the restriction repertoire of the T cells. In contrast to results obtained with other methods of restriction element analysis, we find that approximately 40% of the T-cell response to several antigens is restricted by non-DR class II molecules. Both mutational analysis and blocking by class II specific monoclonal antibodies demonstrate that the non-DR restricted responses derive from DQ and DP-encoded determinants. We also find specific DR/DQ haplotype preferences for the presentation of some but not all antigens. Using a mutant that expresses only the DQ1 molecule, and is derived from a DR1, DQ1 parent line, we demonstrate a functional split of serologically defined DQ1 molecules consistent with the electrophoretic variation reported between DQ1 molecules linked to DR1 and those linked to DR2. Pairs of mutants that differ by expression of a single class II protein reveal a much broader use of available class II restriction elements than previously recognized.

Antibodies, Monoclonal↗

PubMed1990

Identification of cross-reactive T cell restriction epitopes located on the DR7 beta 1 and DR beta 4 molecules.

L cell fibroblasts transfected with HLA class II cDNA clones isolated from a cDNA library produced from a DR7 homozygous cell line were used as antigen-presenting cells (APC) for three HLA DR-restricted, diphtheria toxoid-specific T-cell clones in order to assess the antigen-presenting ability of the transfectants and to define the class II restriction of each clone. Class II-expressing transfectants are capable of presenting antigen to antigen-specific T-cell clones, although the transfectants are less efficient at antigen presentation than conventional APC. Paraformaldehyde fixation of transfectants prior to antigen pulsing abrogated antigen presentation, demonstrating that the transfectants require antigen processing. Antigen presentation by transfectants is completely inhibited by CD4-specific monoclonal antibodies (mAb) and one of four DR-specific mAb, whereas antigen presentation by conventional APC is only partially inhibited. Both the DR alpha:DR7 beta 1 and DR alpha:DR beta 4 (DR omega 53) molecules of the DR7 allotype serve as restriction elements for the diphtheria toxoid-specific T-cell clones. One clone is restricted by the DR7 beta 1 molecule, another clone by the DR beta 4(DR omega 53) molecule, and a third clone by a cross-reactive T cell epitope on DR7 beta 1 and DR beta 4(DR omega 53) molecules. The two DR beta 4(DR omega 53)-restricted clones react, however, differently with a panel of HLA-DR DR omega 53-positive human peripheral blood lymphocytes used as APC. Therefore the data presented here clearly document that the DR beta 4 (DR omega 53) chain may serve as restriction elements for DT-specific T-cell clones. They also provide the first evidence for functional cross-reactivity of the products of two different DR beta loci and in addition emphasize the high complexity of the supertypic HLA-DR omega 53 specificity.

Amino Acid Sequence↗

PubMed1985

Physiological responses of the adult male collared peccary, Tayassu tajacu (Tayassuidae), to severe dietary restriction.

Metabolic and hormonal responses of eight adult male collared peccaries (Tayassu tajacu) to an ad libitum diet intake, or 25% of an ad libitum intake, were examined. Blood samples for hematological, serum-biochemical and hormonal profiles were collected at three week intervals during the nine week experiment starting 4 August 1983. Males fed on the restricted diet lost an average of 26% of their body weight during the trial, compared to a slight weight gain for those fed ad libitum. Characteristics of the red and white blood cell populations were not influenced by diet intake, with the exception of mean corpuscular volume, which was consistently lower amongst males fed on the restricted diet. Restricted food intake resulted in significantly elevated serum values for urea nitrogen, urea nitrogen:creatinine, urea index, alpha globulin:beta globulin, gamma globulin:albumin, nonesterified fatty acids, alkaline phosphatase and lactate dehydrogenase isozymes (LD1 and LD2). Restricted food intake resulted in significantly lowered serum values for total alpha globulin, alpha-1 globulin, total beta globulin, beta-1 globulin, beta-2 globulin, glucose, triglycerides, calcium, magnesium, sodium, chloride, copper and triiodothyronine. Serum levels of creatinine, total protein, albumin, alpha-2 globulin, uric acid, total bilirubin, cholesterol, aspartate aminotransferase, alanine aminotransferase, gamma glutamyltransferase, lactate dehydrogenase, phosphorus, calcium:phosphorus, potassium, iron, zinc and thyroxine were unaffected by diet intake level. Semen evaluation indicated spermatogenesis was not affected by dietary restriction despite reductions in scrotal circumference and ejaculate gel volume. Serum testosterone levels were significantly lower among males fed on the restricted diet after nine weeks. These data suggest male libido might be depressed during poor range conditions, while maintenance of spermatogenesis might permit them to take immediate advantage of improved range conditions. Blood analysis of metabolic and hormonal function can provide useful information for predicting the adult male's nutritional and reproductive condition.

Animals↗

PubMed1994

Influence of dietary restriction and soyabean supplementation on the growth of a murine lymphoma and host immune function.

A comparative study was made to reveal the influence of caloric restriction with or without soyabean in the diet on the growth of a murine lymphoma, host survival, serum profile of vitamin A and E and immune status of the host. Caloric restriction delayed and inhibited tumour growth and improved host survival; inclusion of soyabean during restriction enhanced this effect. Dietary restriction both in the absence and presence of soyabean improved the proliferative response of peripheral blood lymphocytes. This was accompanied by increased cytolytic activity of peritoneal macrophages and elevation in serum immunoglobulins (IgG and IgM). Levels of vitamins A and E, which is found to be low in tumour bearing animals, decreased further when maintained in the restricted diet without soyabean, but was raised to normal levels following addition of soyabean in the diet. These observations imply that tumour growth is arrested possibly by insufficient nutrition available due to dietary restriction, for actively proliferating tumour cells and by improvement in host immune mechanism in the presence of soyabean in the diet.

Animals↗

PubMed1992

Effect of caloric restriction on hepatic nuclear DNA damage in male Fischer 344 rats treated with aflatoxin B1.

Caloric restriction is known to reduce chemically-induced tumor incidence in laboratory animals. The effect is believed to be mediated in part by modification of hepatic drug metabolism, including both phase I and phase II enzymes. Using aflatoxin B1 (AFB1) as a model carcinogen, we studied the effect of caloric restriction on the modification of rat liver nuclear DNA by AFB1 and DNA damage due to the formation of apurinic sites from the AFB1-DNA adduct removal process. Caloric restriction reduced the metabolic activation of AFB1 which resulted in a decrease of AFB1-DNA binding by more than 50%. The results of the study of the effect of caloric restriction on the AFB1-induced DNA strand breakage assayed by the alkaline unwinding technique showed that caloric restriction protected the formation of apurinic sites from the AFB1-DNA adducts and reduced the double strand DNA breakages by 1.3-2.5-fold. Thus, the lower initial AFB1-DNA binding and less DNA damage, presumably by the less apurinic sites formed during the depurination process of AFB1-DNA adducts, contributed to the protective effect of caloric restriction.

Aflatoxin B1↗

PubMed1983

Liver polyploidy: influence of age and of dietary restriction.

Polyploidy increases with age in mammalian liver. Since dietary restriction increases lifespan, the question posed in this study was whether liver polyploidy would be influenced by dietary protein restriction. Restricted mice were fed a 4% protein diet, while control mice received a 26% protein diet. Polyploidy was determined from measurements of nuclear diameter on liver of 3 week, 1 1/2, 3, 10, 20 and 24-1/2 month old Swiss albino mice. The percentage of polyploid cells increased steadily with age. The rate of increase, expressed as the polyploidization index, was greater in the fully fed control animals as compared to the animals on protein restriction. Protein restriction thus retards the age associated process of polyploidization of liver nuclei. Nuclear diameter increased slightly with age, but was not influenced by protein restriction.

Aging↗

PubMed1992

Lymphoproliferative responses in diet-restricted and aging Sprague-Dawley rats.

Immunological effects of aging and dietary restriction (i.e., chronic underfeeding without malnutrition) were investigated in male CFY Sprague-Dawley rats. From weaning, diet-restricted animals were given the amount of food that maintained their body weights at approximately 50% of age-matched ad libitum controls. Cells from the spleen, Peyer's patches, brachial/axillary lymph nodes, and mesenteric nodes of 12- and 20-month-old rats were tested in vitro against the T-cell mitogens phytohaemagglutinin and concanavalin A, and the B-cell mitogen Salmonella minnesota lipopolysaccharide (with dextran sulphate). No clear boosting effect of dietary restriction nor age-related decline in T- or B-cell response was observed when a standard foetal bovine serum supplement was used. In general however, serum from diet-restricted rats supported better proliferative responses than serum from age-matched controls, suggesting that dietary restriction may promote lymphocyte proliferation by an indirect mechanism. A feature of this study was the variation in phenotype and immune responsiveness among closely related animals. Thus in an outbred population, any beneficial effects of dietary restriction upon immune responses could be outweighed by variation among individuals.

Aging↗

PubMedDate not supplied

Fatty acid composition of adipose tissue in aged rats: effects of dietary restriction and exercise.

Little is known about the long-term effects of diet restriction, exercise, and aging on the fatty acid composition of adipose tissue. To investigate these effects, male Wistar rats were placed in one of four experimental groups: 1) ad libitum fed, nonexercised; 2) ad libitum fed, exercised; 3) diet restriction, nonexercised; and 4) diet restriction, exercised. Exercise consisted of swimming on alternate days, and diet-restricted animals were fed every other day. Animals were assigned to one of the four treatment groups at weaning, and the fatty acid composition in the epididymal, perirenal, and inguinal fat depots were determined every 4 months from 12 to 28 months of age. All animals were fed the same diet so the relative percentage of fatty acids in the diet did not contribute to the changes observed in adipose tissue fatty acid composition. Regardless of treatment groups and fat depot, the concentration of saturated fatty acids in adipose tissue tended to decrease as body weight increased. Saturated fatty acid composition was greater in the diet-restricted than the ad libitum-fed groups. The saturated fatty acid concentration was different between the three fat depots (perirenal 27.8%, inguinal 24.7%, and epididymal 21.6%; P < 0.001). The linoleic acid content did not change with age in the epididymal or perirenal fat depot; however, the change was significant for the inguinal fat depot. Following a large decrease in inguinal linoleic concentration through 20 months, linoleic acid concentration increased in all treatments between 20 and 28 months. The results indicate that diet restriction, exercise, and aging have significant effects on adipose tissue fatty acid composition and these effects are different depending on the anatomical location of the adipose tissue depot.

Adipose Tissue↗

PubMed1994

Incidence of apparent restricted diffusion in three different models of cerebral infarction.

High speed magnetic resonance imaging (MRI) and short diffusion times are used to investigate the appearance of restricted diffusion in three different models of cerebral infarction. The models are: the middle cerebral artery occlusion (MCAO) model in the rat, the carotid occlusion model in the gerbil, and the Rose Bengal microvascular occlusion model in the rat. All three were investigated for 16 b-values equally spaced between 10 and 1510 s/mm2 using two distinct experiments. In the ct (constant time) experiment, the diffusion time was held constant at 11.7 ms while the b-value was varied with the gradient strength. In the cg (constant gradient) experiment, the gradient strength was held constant and the b-value increased by varying the diffusion time from 4.4 to 11.7 ms. A monoexponential decay of the signal intensity with b-value in the ct experiment accompanied by nonmonoexponential (NME) decay in the cg experiment is indicative of restricted diffusion. As this phenomenon is detectable only at short diffusion times, it cannot be due to restriction by impermeable membranes, and we have thus termed this apparent restriction. For the MCAO model and the carotid occlusion model, apparent restriction was found both inside the infarct territory and in some regions outside it. No definite evidence for restriction was found for the Rose Bengal model, which was, however, only studied from 24 h post-insult.

Animals↗