Induction of tolerance to an experimental cardiac allograft through intrathymic inoculation of class II major histocompatibility complex disparate antigens.

Indefinite donor-specific tolerance to a cardiac allograft can be induced through pretransplantation intrathymic injection of donor spleen cells and a single intraperitoneal injection of antilymphocyte serum. This study was designed to determine whether this phenomenon was reproducible with grafts differing in either class I major histocompatibility complex only or class II MHC only. Donors of cells and hearts in all experiments were RP rats. Class I MHC disparate grafts were performed by placing an RP heart into a Lewis recipient, and class II disparate grafts were performed with RP donors and Wistar Furth recipients. Lewis (n = 10) and Wistar Furth (n = 10) recipients underwent intraperitoneal injection of 1 ml antilympocyte serum and intrathymic injection of 5 x 10(7) RP spleen cells. Three weeks later, heterotopic cardiac transplantation was done with a heart from an RP rat. Control rats had no pretreatment or received antilympocyte serum alone. Without pretreatment, RP hearts survived 7 to 9 days (mean 8 days) in Lewis recipients (n = 5) and 9 to 14 days (mean 12 days) in Wistar Furth recipients (n = 5). Antilymphocyte serum alone produced slight prolongation of graft survival. Lewis rats pretreated with class I disparate RP splenocytes and antilympocyte serum had graft survivals of 8 to 27 days (mean 14 days), not significantly different from the results with antilympocyte serum alone. Class II disparate RP grafts placed in pretreated Wistar Furth rats had significant prolongation of graft survival, with four of five grafts surviving longer than 60 days (p < 0.01 vs antilympocyte serum alone). These results suggest that a disparity at the class II locus of the major histocompatibility complex is critical for the induction of cardiac allograft tolerance after intrathymic inoculation of allogeneic cells.

Different immune effects on cardiac allografts and xenografts induced by neonatal intrathymic inoculation with allogeneic and xenogeneic antigens.

BACKGROUND: In a previous study we demonstrated that intrathymic exposure of neonatal rats to both alloantigens and xenoantigens produced tolerance only to subsequent cardiac allografts and not to xenografts implanted when the animals were 6 weeks old. Interestingly, graft recipients were not sensitized to the xenografts as observed in the adult model. This study was designed to investigate whether earlier grafting would produce tolerance to cardiac xenografts in animals pretreated by neonatal intrathymic inoculation with allogeneic and xenogeneic cells. METHODS: All recipient animals were Lewis rats. Donors were either Lewis Brown Norway rats or Golden Syrian hamsters. Lewis Brown Norway rat and hamster splenocytes (25 x 10(6) cells in a volume of 0.01 ml) were inoculated percutaneously into the thymus of neonatal recipients (n = 22). At age 4 weeks, five pretreated recipients underwent cervical heterotopic heart transplantation with rat hearts, and 2 weeks later abdominal heterotopic transplantation was done with hamster donors. A second group ( n = 6) received hamster hearts as the first graft and then grafts from rat donors. The third group underwent rat followed by hamster heart transplantation at age 6 to 7 weeks. RESULTS: Mean rat allograft survival time for groups 1,2, and 3, respectively was 49.8 days with 4 of 5 surviving indefinitely, 4.3 days with 2 of 3 surviving indefinitely, and 42 days with 7 of 11 surviving indefinitely (p < 0.01 versus untreated control animals, 7.8 days, n = 5). In rats undergoing transplantation at 4 weeks, cardiac xenografts were rejected in 2.5 days 2.3 days, which was significantly shorter than xenograft survival (3.3 days) in rats that underwent transplantation at an older age (p < 0.02), in naive rats (p < 0.05), and in rats treated with hamster cells alone (p < 0.05). Mixed lymphocyte reaction showed a diminished proliferative response to Lewis Brown Norway rat cells in pretreated rats, which retained the ability to respond in culture to hamster cells (p < 0.05). CONCLUSIONS: Earlier grafting in rats pretreated as neonates produces allograft tolerance but may accelerate rejection of xenografts. Preliminary mixed lymphocyte reaction results suggest that only the alloimmune cellular proliferative response is abrogated by this pretreatment.

Tolerance to cardiac allografts requires a time lag between intrathymic treatment and transplantation.

Permanent tolerance to an experimental heterotopic cardiac allograft can be achieved by pretreatment with antilymphocyte serum (ALS) and intrathymic inoculation of donor cells. Most successful experimental protocols have employed a time lag of 2 to 3 weeks between intrathymic pretreatment and transplantation, which makes this treatment strategy impractical for clinical heart transplantation. In these experiments we modified the standard protocol by giving ALS 24 hr prior to both intrathymic injection of donor cells and heterotopic transplantation. Seven Lewis rats had intraperitoneal injection of 1 ml of ALS and 24 hr later underwent intrathymic injection of 5 X 10(7) donor Lewis-Brown Norway (LBN) splenocytes and heterotopic cardiac transplantation using an LBN donor. Mean graft survival was 24.4 days, significantly longer than the 7.8-day graft survival observed in untreated Lewis recipients (n = 5) (P < 0.02). However, graft survival was not different from that observed in Lewis rats pretreated with ALS alone (n = 5) (25.8 days, P = NS). Permanent graft survival was produced in two rats receiving only A-LS and in one rat receiving both ALS and intrathymic inoculation. In these experiments it appears that prolongation of graft survival may have been due to the effect of A-LS alone. These results suggest that there is a critical time period between intrathymic inoculation and transplantation that is needed for permanent tolerance to be induced consistently. This may be due to the kinetics of the effects of ALS on alloreactive T-lymphocytes or to a time-dependent requirement for antigen processing in the thymus.

Laser trabeculoplasty.

Over a decade, laser trabeculoplasty has evolved from being a novel new treatment to one that is a commonly accepted intervention in the management of open-angle glaucoma. Despite its widespread use, however, there are still many unanswered questions about laser trabeculoplasty, including its mechanism of action and the ideal treatment parameters. In this review, we will discuss the history of the technique, the clinical experience, and some of the experimental studies that have been conducted to answer the questions regarding its mechanism of action.

Is the high concentration of ascorbic acid in the eye an adaptation to intense solar irradiation?

Ascorbic acid is known to exist in high concentration in the aqueous humor of the eye in many species. It has been observed that diurnal mammals have a very high concentration in aqueous humor whereas nocturnal mammals do not. It has been hypothesized that ascorbic acid protects the eye from the harmful effects of sunlight. We have discovered that of two closely related species of spiny mice, the diurnal species (Acomys russatus) has a concentration in aqueous humor that is 35 times higher than that of the nocturnal species (Acomys cahirinus). Studies of these two species may be fruitful to extend what is known about adaptation of the eye to protect itself from intense solar radiation.

Infectious crystalline keratopathy.

A 57-year-old white female was noted to have unusual intrastromal crystalline-like opacities in her failing right corneal graft five months after transplant surgery. Cultures grew Streptococcus viridans; since the graft was failing it was replaced and sent for pathologic examination. H&E stains revealed intrastromal pockets of basophilic material between the lamellae; there was a notable absence of inflammatory cells. Gram's stain showed the basophilic material to be gram positive and transmission electron microscopy confirmed the presence of intrastromal pockets of bacteria. The term "infectious crystalline keratopathy" was coined by Meisler for this entity, which occurs following longterm topical steroid usage. The quite clinical appearance may make its infectious etiology unapparent. Management consists of discontinuation of the steroids and the administration of antibiotics; continued infection, corneal scars, or graft failure may occur and transplantation is then required.

Ascorbic acid levels in the aqueous humor of nocturnal and diurnal mammals.

Ascorbic acid concentration is known to be very high in the aqueous humor of humans and most animals. The role it might play in ocular function is a subject of conjecture. Some have proposed that it might protect the eye against light-induced damage. We examined the aqueous humor from 22 species of mammals to determine the range of levels and to see if there was a correlation with behavior. A wide range of ascorbic acid levels in the aqueous humor was found. Most of the animals considered to be diurnal had higher ascorbic acid levels than the nocturnal animals. This would suggest that ascorbic acid in the aqueous humor may play a protective role in those animals who are most exposed to light. Regardless, any theory proposing a role for ascorbic acid in the eye in mammals must take the wide range of ascorbic acid levels into account.

Aqueous humor flow during sleep.

The rate of aqueous humor flow of 19 normal subjects was measured by a fluorescein clearance technique during the day, during sleep (at night) and during sleep deprivation (at night). Subjects engaged in routine activities during the daytime measurements and slept in comfortable quarters during the nighttime measurement. They remained awake and active during the sleep deprivation study. The nomographic method of Coakes and Brubaker , a method in which the subject is not disturbed at all during the critical 5-hr period, was used to calculate flow. Flow was lower during sleep in all but one subject. The range of nighttime flow suppression was 8% to 68% in all of the other subjects with a mean suppression of 45% +/- 20%. Aqueous flow was lower at night even when the subjects were awake but was lowest during sleep at night. The reduction of aqueous flow during sleep is comparable with the suppression that can be achieved with carbonic anhydrase inhibitors or beta adrenergic blockers.

The mechanism of betaxolol, a new ocular hypotensive agent.

A single drop, double-masked study was performed using 1% betaxolol ophthalmic solution in 24 normal subjects. Subjects received one drop of this new drug to one eye and a placebo drop to the fellow eye. Aqueous humor flow was measured using the technique of fluorophotometry, and outflow resistance was measured using tonography. Betaxolol suppressed aqueous humor flow in all individuals. The range of suppression was from 7 to 51%, with a mean suppression of 32 +/- 13%. The drug had no consistent effect on outflow resistance, and no pupillary changes were seen.