Implications/Analysis of Family History

FD’s mother is a hepatitis B virus (HBV) carrier who was identified during maternal screening and was found to have anti-HBc and anti-HBe IgG antibodies specific for the HBV core and HBe (pre core) antigens (HBcAg and HBeAg, respectively). HBeAg is associated with HBcAg (Fig. 39-1). Additional enzyme-linked immunosorbent assays (ELISAs) revealed that the mother was positive for the HBs antigen (HBsAg) but negative for HBeAg. Had the mother been HBeAg positive, the infant would have been given gamma globulin (anti-HBV antibodies) and the first of the three required HBV vaccine antigens. Because the mother was negative for HBeAg, the child had not been given either therapy, according to the regulations in place. HBsAg may also be detected in liver biopsies (Fig. 39-2).

FIGURE 39-1 Serologic and clinical patterns observed during acute and chronic hepatitis B infection.

(From Murray P, Rosenthal K, Kobayashi Y, Pfaller M: Medical Microbiology, 4th ed. St Louis, Mosby, 2002; redrawn from Hoofnagle JH: Annu Rev Med 32:1–11, 1981.)

FIGURE 39-2 Negative-stain electron micrograph of plasma from a patient with acute hepatitis B virus infection. The complete virus (Dane) particles (d) have a lipid envelope with a loop containing the double-stranded DNA genome in a cuboidal nucleocapsid. The tubular (t) and vesicular (v) particles comprise only viral lipid envelope. (bar =100 nm.)

(From Hart CA, Shears P: Color Atlas of Medical Microbiology, 2nd ed. St Louis, Mosby, 2004.)

In some regions, however, different regulations are in place and infants born to mothers who are HBsAg positive are immunized at birth. Otherwise, the first dose of the HBV vaccine is deferred until the infant is 2 to 6 months of age. There are reported cases when vertical transmission has occurred in the first months post partum if the mother becomes infected during this time or if she is a chronic carrier. Additionally, development of FHF in infants, despite appropriate prophylaxis, has been documented.

The mother had been prescribed interferon alfa (IFNα) and a nucleoside analog. Both of these therapeutic agents have been approved by the U.S. Food and Drug Administration (FDA) for treatment of chronic EBV infection.

Analysis/Implications of Clinical History

For many children the etiology of FHF cannot be identified; however, common causes include viruses, toxins, congenital abnormalities (e.g., biliary atresia), metabolic diseases, and hepatotoxic drugs (e.g., acetaminophen overdose). Included in the list of viruses that can cause FHF are herpes simplex virus, adenovirus, cytomegalovirus, and EBV. Hepatitis A and hepatitis B are common causes of FHF in children and infants, but hepatitis C is not. Patients with hepatitis non–A-E infections and FHF have a higher fatality rate. Symptoms of severe liver failure include jaundice, fever, bleeding, abdominal distention and pain, fatigue, and vomiting.

Implications/Analysis of Laboratory Tests

The infant’s blood cell count indicated a leukocytosis (see Appendix for normal reference values) and thrombocytopenia. Biochemical tests revealed high serum bilirubin, increased ALT and AST levels, decreased albumin, an increased INR (>4; normal ˜1.0), and an increase in PT (normal: 12 to 14 seconds) and aPTT (normal: 25 to 38 seconds). Overall, the results of these tests indicated severe liver damage. Patients with severe FHF have an increased PT because the liver makes the majority of the clotting factors. Consequently, bleeding is common. A diagnosis of HBV infection was confirmed when the ELISA revealed anti-HBc IgM antibodies in FD’s serum, as well as anti-HBc IgG antibodies, transferred via the placenta ELISA. On the basis of the results of these tests, abdominal CT and ultrasonography were requested.

Note: The PT refers to the time required for thrombin to convert plasma fibrinogen to fibrin. The aPTT measures the clotting time of plasma as a result of the activation by factor XII after serum contact with a negatively charged surface (e.g., silica).

Outcome of the Xenograft

The porcine liver transplant itself goes well. There is, in fact, no evidence of early graft loss (hyperacute rejection, see later). The team is also aware of potential problems with the so-called delayed hyperacute rejection response, occurring several days later and also believed to be associated with antibody-mediated immune activation, this time at the (graft) endothelial bed, resulting again in thrombosis and graft failure. Once again this, too, seems not to play any part in the events occurring postoperatively for this transplant. The anti-HBc IgM antibody titer does fall in the early days post transplant, which it was hypothesized may have reflected the failure of pig liver to support HBV replication.

Challenges of Xenotransplants: Hyperacute Rejection

Transplantation across species barriers (xenotransplantation) is fraught with risks not seen in allotransplants. The first problem that arises from major species barriers is a ubiquitously expressed carbohydrate structure Galα1-3-Galβ1-4GlcNAc-R (α-galactosyl epitope). Humans and Old World monkeys do not express in their genome the gene encoding α1-3-galactosyltransferase (α1-3GT), an enzyme that synthesizes α-galactosyl epitopes on glycoproteins and glycolipids (using uridine diphosphate galactose [UDP-Gal]). Hence we, unlike multiple other species, including pigs, cats, dogs, horses, and New World primates, do not link Galα1-3 to the ends of carbohydrates expressed at the surface of cells. Accordingly, lacking tolerance to this α-galactosyl epitope, we have abundant antibodies (anti-Gal) that target this particular epitope (induced in response presumably to exposure from commensal bacteria, expressing this on their cell walls). Transplantation of organs expressing the α-galactosyl epitope on the surface leads to rapid antibody deposition at the cell surface, complement fixation, and thrombosis, with graft loss. Consequently, it has been thought that if complement activation could be controlled, this form of rejection (hyperacute rejection) would be minimized.

Zoonosis: Increased Risk of Porcine Endogenous Retrovirus Infection with DAF Pigs

There is evidence that any time cells from different species are mixed a zoonotic infection is a potential problem. Consequently, the use of porcine tissues for xenografts also increases the potential for the transmission of PERV. Although heterografts using pig aortic heart valves have been used for years, these valves were sewn into a cloth-covered frame and therefore somewhat inaccessible to the immune system. Additionally, the valve recipients were not immunosuppressed.

As described earlier, one of the major problems with the use of porcine tissues has been hyperacute rejection because these tissues express α-galactosyl epitope to which humans have natural antibodies (anti-Gal). The presence of DAF proteins within the membranes of porcine cells should disrupt the formation of C3 convertase enzymatic complexes on the cell surface. These complexes form after activation of complement—in this case, by anti-Gal antibody-α-galactosyl epitope immune complexes.

Attempts to eliminate this immunologic reaction, however, may contribute to the transmission of PERV because the DAF proteins will be incorporated into the viral envelope when the virus buds from the DAF pig cells. Consequently, the PERV will be as resistant to complement-mediated destruction after formation of immune complexes on their envelopes as the porcine cells.

Note: In vitro studies have shown that human anti-Gal antibodies inactivate retroviruses released from animal cells by triggering the activation of the classical pathway of complement.

Other Approaches That Also Increase Risk of PERV

In another approach, the gene that encodes the enzyme α1-3GT has been knocked out and so the α-galactosyl epitope is not expressed in the porcine tissues. The absence of this epitope in these tissues would eliminate the hyperacute rejection problem. However, in the transgenic porcine infected cells, the PERVs that bud from these cells would not express this epitope either. Therefore, the anti-Gal antibodies would not be able to target these viral particles, thereby eliminating what may be an effective mechanism for viral elimination. The extent to which complement plays a role in viral immunity in vivo is not clearly defined. However, in vitro studies have shown that anti-Gal antibodies inactivate retroviruses released from animal cells. The same argument has been used for the removal of human anti-Gal antibodies from the transplant recipient.

Other Scenarios for Zoonosis

The potential for zoonotic infection is not limited to xenografts. In many in vitro fertilization (IVF) centers, human eggs and preimplantation embryos are cultured on animal feeder cells, most commonly bovine kidney cells, before reimplantation to the mother. Whether this poses a long-term hazard is not known. To eliminate the risk of zoonosis, some IVF centers use only cells derived from one of the embryo’s biologic parents.