Etiology and Pathogenesis

Broadly defined, HUS and TTP are syndromes whose pathologic correlate is thrombotic microangiopathy (TMA). TMA describes the microvascular occlusion that occurs most frequently within capillaries and arterioles (Figure 64-1). It is seen histologically as thrombi, endothelial cell swelling, luminal narrowing, and fibrinoid necrosis of the vessel wall. TMA results from dysfunction of the endothelial cell–platelet interface and can originate from various underlying mechanisms, which form the basis for an etiologic classification. HUS can result from infectious causes, genetic causes, or medication-related causes or in association with secondary discrete pathologic entities. TTP results from a deficiency of von Willebrand factor-cleaving protease, (vWF-cp), which can be either acquired because of the presence of an autoantibody or congenital resulting from a mutation in the ADAMTS13 gene.

Figure 64-1 Thrombotic disorders: hemolytic-uremic syndrome.

The majority of childhood cases of HUS occur after a prodromal diarrheal illness. For this reason, it has been termed D+ or “typical” HUS, in contrast to the less common forms of HUS not associated with a diarrheal prodrome, known as D- or “atypical” HUS. A more precise classification for D+ HUS is Shiga toxin–associated HUS (Stx HUS), which is known to cause 90% of all HUS cases in children. As the name suggests, Stx HUS occurs as a result of Shiga toxin–producing organisms. The most common of these is Shiga toxin–producing Escherichia coli (STEC), also known as verocytotoxin-producing E. coli, so-called for their ability to lyse vero cells, a primate kidney cell line with epithelial characteristics. And of these, the most common serotype is O157 : H7, which expresses somatic (O) antigen 157 and flagellar (H) antigen 7. Stx HUS can occur at any age but primarily affects children younger than 5 years of age; the peak incidence is between the ages of 6 months and 4 years. It occurs both sporadically and in the form of epidemic outbreaks, most commonly during the summer and autumn months and largely in rural areas. The disease has an annual incidence of two to three per 100,000 children younger than 5 years of age in North America and Western Europe. The incidence decreases among older children. In countries such as Uruguay and Argentina, STEC infections are endemic and cause HUS in about 10.5 per 100,000 children per year.

The primary reservoir for STEC is cattle. Environmental sources of the infection include undercooked beef or poultry, deer jerky, unpasteurized milk or other dairy products, unpasteurized apple cider, fruits, vegetables, and contaminated municipal or swimming water. Additionally, STEC infection can be acquired as zoonoses from petting zoos, from human-to-human contact, or from urinary tract infection with the organism. Testing of stool may confirm Stx-producing organisms in up to two-thirds of cases, but the environmental source is rarely discovered in sporadic cases.

HUS not associated with enteropathic STEC (NStx) encompasses a disease group with heterogeneous etiologies. NStx HUS accounts for about 10% of all cases of HUS in children. Of this group, the most common cause is Streptococcus pneumoniae-associated HUS (pneumococcal HUS), accounting for about 40% of children with NStx HUS. Pneumococcal HUS has an estimated incidence of 0.4% to 0.6%, although the possibility that this underestimates the true incidence is noted by an overall lack of recognition of the disease.

Shiga Toxin–Associated Hemolytic-Uremic Syndrome

The most common causative organism of Stx-HUS is enterohemorrhagic E. coli (EHEC). Among the EHEC that cause HUS, serotype O157 : H7 is predominant, accounting for 70% of cases in North America and Western Europe. There are several known STEC that are non-O157 strains (e.g., O111 : H8, O103 : H2, O121, O145, O26, and O113). Shigella dysenteriae type 1-associated HUS occurs more frequently in developing countries in Asia and Africa but rarely in industrialized countries. Other organisms such as Aeromonas spp. and Citrobacter freundii have been known to cause Stx HUS. E. coli O157 : H7 infection can be confirmed by plating a stool sample on sorbitol-MacConkey agar. The O157 : H7 strains cannot ferment sorbitol during an overnight incubation and appear as translucent colonies. The organism can be further confirmed by rapid assays that allow detection of Shiga toxin in the stool, including non-O157 serotypes. It is generally recommended to test stool samples by sorbitol-MacConkey agar as a first screening. The potential to detect the organism is higher in the first 6 days after onset of diarrhea.

Shiga toxin is a potent exotoxin that functions as the virulence factor in STEC. All members of the Stx family share some degree of sequence homology, the protein products of which have common structures that include an enzymatically active A subunit linked to a pentameric B subunit (A₁B₅). The STEC can elaborate at least four plasmid-encoded Shiga-toxins: Stx1, Stx2, Stx2c, and Stx2d (Stx is used interchangeably with the term VT). Stx1 is nearly identical to the classic Shiga toxin, and Stx2 shares 60% homology with classic Shiga toxin. The Stx may be expressed individually or in combination with two or three different Shiga-toxins. The Stx2 gene is carried by most E. coli O157 : H7, and its expression individually causes more severe disease than Stx1 or the combination of Stx1 and Stx2.

Stx is responsible for the endothelial toxicity of HUS-causing STEC, giving rise to the pathologic hallmark of TMA. It is produced in the bowel and translocated into circulation, where it can localize to the glomeruli, gastrointestinal tract, pancreas, and various other host tissues by a mechanism that has yet to be fully understood. Central to enterocyte entry and subsequent toxemia, the Stx B subunit binds to a cell surface terminal carbohydrate moiety of the globotriaosylceramide receptor (Gb3). The Gb3 receptor is a key determinant for cell sensitivity to Stx, and along with enterocytes and other cell types is present on glomerular endothelial cells, thereby targeting the toxin to the renal microvasculature. In the intestine, binding of Stx to Gb3 commences a sequence of events beginning with receptor-mediated endocytosis. Stx can follow several different pathways: (1) it can be delivered intact to the intestinal submucosa and circulation via transcytosis; (2) it can induce direct cytotoxicity by trafficking to the cell endoplasmic reticulum via the Golgi apparatus in a process known as retrograde transport; or (3) it can, in lower concentrations, alter gene and protein expression of the cell without inducing cell death. Transcytosis of toxin to the intestinal microvascular circulation is thought to give rise to the characteristic intestinal lesion that has the clinical–pathologic manifestation of bowel wall edema, thrombosis, and hemorrhage. Neutrophils localize to the intestinal mucosa during STEC infection, where they are thought to transport Stx to extraintestinal sites. They are known to bind Stx by a distinct receptor with a lower affinity than Gb3. The toxin is therefore not endocytosed, which allows it to be freely unloaded at various target sites (Figure 64-2).

Figure 64-2 Effects of Shiga toxin on enteric cells and endothelial cells.

The cytopathic effects of Stx result when the A subunit becomes enzymatically active within the host cell by proteolytic cleavage and in turn cleaves its target adenine residue (A4324) on the 28S rRNA of the 60S ribosomal subunit. This action blocks binding of the aminoacyl-tRNA to the subunit with resultant protein synthesis inhibition and cell death. Alternatively, subinhibitory concentrations of toxin are known to alter gene expression in the host cells. Stx has been found to increase expression of prothrombotic and inflammatory genes that affect the properties of endothelial cells. Various cell types undergo toxin-mediated increase in cytokine and chemokine production. For example, subinhibitory toxin concentrations cause endothelial cells to upregulate interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) as well as endothelin-1 and tissue factor. The effector functions of these molecules result in leukocyte migration or adhesion, vasoconstriction, and a procoagulant cellular milieu. Additionally, monocytes increase cytokine production of tumor necrosis factor-α and IL-1β, factors known to sensitize host cells to toxin via upregulation of Gb3.

A major virulence cofactor of pathogenic STEC is the ability of the organism to exploit the host enterocyte by secreting its own bacterial receptor into the cell such that it is expressed on its apical surface, in turn allowing firm attachment of the organism along the intestinal mucosal surface. This receptor incorporation occurs through a macromolecular complex called the type 3 secretion system, which results in cytoskeletal changes with associated loss of normal villous architecture, giving rise to a characteristic “attaching and effacing” lesion on the host cell. The genetic locus responsible for this process is found on what are known as pathogenicity islands of the bacterial chromosome. The genes encode the machinery of the type 3 secretion system as well as intimin, expressed on the bacterial cell surface, and the intimin receptor, which is translocated into the host cell for surface expression.