Infection of cultured human pancreatic B cells with reovirus type 3

Abstract

Summary. The capacity of reovirus type 3 to infect insulin-producing B cells was studied in human pancreatic cell cultures. Antibody to reovirus was labelled with fluorescein isothiocyanate and antibody to insulin was labelled with tetramethyl rhodamine isothiocyanate. By using a double-labelled immunofluorescent antibody technique, it was shown that only about 6% of the insulin-containing human B cells in culture became infected when inoculated with unpassaged reovirus type 3. However, by repeated passage of the virus in human pancreatic B cell cultures, the percentage of infected B cells increased to 27%, and the virus titre in cultures rose from 8.0 x 104 pfu/ml in the first passage, to 4.9 x 106 pfu/ml in the 5th passage. As measured by radioimmunoassay, the intracellular immunoreactive insulin began to decrease at 24h after infection. This decrease roughly paralleled the increase in virus titre. In contrast, there was relatively little change in immunoreactive insulin in cultures inoculated with unpassaged reovirus type 3. These studies show that the ability of reovirus type 3 to infect human B cells is enhanced by serial passage in human pancreatic cell cultures and that the infection resulted in the destruction of B cells and release of insulin.Key words: Virus-induced diabetes mellitus, human pancreatic B cell cultures, double-labelled immunofluorescent antibody staining, reovirus type 3, virus passage. study, it was shown that a Coxsackie virus B4 variant, isolated from the pancreas of a 10-year-old child who died of diabetic ketoacidosis, produced diabetes in mice by infecting and destroying B ceils [7].It is difficult, however, to demonstrate in vivo that viruses replicate in human B cells and produce diabetes in man. As a practical model, an in vitro system has been developed to determine if viruses are capable of infecting human B cells. By use of a double-labelled immunofluorescent antibody technique, it has been shown that human B cells grown in culture were susceptible to infection by mumps [8], Coxsackie virus B3 [9], and Coxsackie virus B4 [7].We have previously shown that reovirus type 3, passaged in murine pancreatic B cell cultures, produced an insulitis when inoculated into 1 to 2 week old mice [10]. In the present investigation, the double-labelled immunofluorescent antibody technique has been used to determine whether or not human B ceils grown in culture were permissive to reovirus type 3, and whether the capacity of the virus to infect human B cells was enhanced by repeated passage in human B cell cultures. Radioimmunoassay for immunoreactive insulin was used to evaluate the effect of the infection on intracellular and extracellular insulin. Materials and Methods Pancreatic B Cell CulturesThe possibility that viruses might cause some cases of insulin-dependent diabetes (IDD) by infecting and destroying pancreatic B cells has received considerable attention. Of the numerous viruses implicated, mumps and members of the Coxsackie virus B group (particularly B4) have been most often suggest...

Reovirus Infection Enhances Expression of Class I MHC Proteins on Human β-Cell and Rat RINm5F cell

“…Reoviruses 1 and 3 are notable for their ability to induce autoimmune disease, including diabetes, in susceptible mice (20,21). Infection of human p-cells (22) and RINm5F cells (23) in vitro with reovirus results in both functional and cytopathic effects. Our study demonstrates that an additional and novel aspect of reovirus infection of pancreatic p-cells is the stimulation of class I MHC protein expression.…”

“…In this study, we have examined the effects of reovirus 1 and reovirus 3 infection on the expression of MHC proteins on human p-cells and rat insulinoma cells. Reoviruses 1 and 3 are appropriate candidates because they have been shown to induce autoimmunity and diabetes in susceptible mice (20,21) and to infect human (22) and rodent (20,23) (J-cells in vitro.…”

Ammonium inhibits processing and cytotoxicity of reovirus, a nonenveloped virus.

“…Type 1, when injected into newborn mice, infects ependymal tissue and causes hydrocephalus (9), whereas type 3 infects neurons and causes a rapidly fatal encephalitis (10). Both types 1 and 3 reovirus can also infect cells ofthe endocrine pancreas and may cause diabetes (11)(12)(13). Type 1 also produces an autoimmune syndrome associated with growth retardation (14).…”

Specific plasma membrane receptors for reovirus on rat pituitary cells in culture.

“…In mice, type 1 reovirus is known to produce an ependymitis and to infect cells of the endocrine pancreas and anterior pituitary, which leads to the development of autoimmune syndrome associated with diabetes, runting, and the development of antibodies to insulin and growth hormone (9). In vivo type 3 reovirus preferentially infects neurons, but can also infect endocrine cells (10). GH4C1 are a clonal line of rat pituitary cells (11)(12)(13) GH4C1 cells were grown to confluency on 100-mm dishes in Ham's F10 media supplemented with 15% horse serum and 2.5% fetal calf serum.…”