Variant Creutzfeldt-Jakob disease strain is identical in individuals of two PRNP codon 129 genotypes

Abstract

In 2004, a subclinical case of variant Creutzfeldt-Jakob disease in a PRNP 129 methionine/valine heterozygous individual infected via blood transfusion was reported, and we established that the spleen from this individual was infectious. Since host genetics is an important factor in strain modification, the identification of variant Creutzfeldt-Jakob disease infection in a PRNP 129 methionine/valine heterozygous individual has raised the possibility that the properties of the variant Creutzfeldt-Jakob disease agent could change after transmission to this different genetic background and concerns that this could lead to a more virulent strain of variant Creutzfeldt-Jakob disease. The variant Creutzfeldt-Jakob disease strain has to date been characterized only in methionine homozygous individuals, therefore to establish whether the strain characteristics of variant Creutzfeldt-Jakob disease had been modified by the host genotype, spleen material with prion protein deposition from a PRNP 129 methionine/valine individual was inoculated into a panel of wild-type mice. Three passages in mice were undertaken to allow stabilization of the strain characteristics following its passage into mice. In each passage, a combination of clinical signs, neuropathology (transmissible spongiform encephalopathy vacuolation and prion protein deposition) were analysed and biochemical analysis carried out. While some differences were observed at primary and first subpassage, following the second subpassage, strain characteristics in the methionine/valine individual were totally consistent with those of variant Creutzfeldt-Jakob disease transmitted to 129 methionine/methionine individuals thus demonstrated no alteration in strain properties were imposed by passage through the different host genotype. Thus we have demonstrated variant Creutzfeldt-Jakob disease strain properties are not affected by transmission through an individual with the PRNP methionine/valine codon 129 genotype and thus no alteration in virulence should be associated with the different host genotype.

Keywords: PRNP; prion; transmissible spongiform encephalopathy; variant Creutzfeldt-Jakob disease.

Figure 1

Comparison of incubation periods and TSE vacuolation profiles in wild-type mice challenged with brain and spleen homogenates from four cases of vCJD. The four cases used to challenge wild-type mice were: an asymptomatic MV blood recipient (MV^R), the MM donor to that individual (MM^D), an MM blood donor (MM^D2008) and the associated MM blood recipient (MM^R2008). (A) Incubation periods in RIII, VM and C57BL6 mice. Incubation periods were calculated in mice showing clinical and pathological signs of TSE disease. (B) TSE vacuolation profiles in C57BL6 mice (n ≥ 6). (C) TSE vacuolation profiles in VM mice [n ≥ 6 in MM^D (CNS), MM^R2008 and MM^D2008, n = 4 in MM^D (spleen)]. (D) TSE vacuolation profiles in RIII mice (n ≥ 6). All data shows mean ± SEM. Brain region areas: G1–9, grey matter scoring areas; G1, medulla; G2, cerebellum; G3, superior colliculus; G4, hypothalamus; G5, thalamus; G6, hippocampus; G7, septum; G8, retrosplenial and adjacent motor cortex; G9, cingulate and adjacent motor cortex. W1–W3, white matter scoring regions: W1, cerebellar white matter; W2, mesencephalic tegmentum; W3, cerebral peduncle. Dpi = days post inoculation.

Figure 2

Abnormal PrP deposition in wild-type mice challenged with brain and spleen homogenates from two cases of vCJD. The two cases comprised an asymptomatic MV blood recipient (MV^R) and the MM donor to that individual (MM^D CNS and spleen). (A) RIII, (B) C57BL6, and (C) VM, mice challenged with MV^R. (D) RIII, (E) C57BL6, and (F) VM, mice challenged with MM^D (CNS). (G) RIII, (H) C57BL6, and (I) VM, mice challenged with MM^D (Spleen). (J) Abnormal PrP deposition in CA2 region of hippocampus from RIII mouse (inset region of A). (K) Abnormal PrP deposition in CA2 region of hippocampus from C57BL6 mouse (inset region of E). (L) PrP plaque in the corpus callosum of a VM mouse (inset region of F). Scale bars = 200 µm (A–I); 100 µm (J–L).

Figure 3

Incubation periods in wild-type (RIII, C57BL6 and VM) mice from the first and second mouse subpassage of brain and spleen homogenates from four cases of vCJD. The four cases comprise: an asymptomatic MV blood recipient (MV^R), the MM donor to that individual (MM^D), an MM blood donor (MM^D2008) and the associated MM blood recipient (MM^R2008). Incubation periods were calculated in mice showing clinical and pathological signs of TSE disease. Black box indicates significant changes in incubation periods (P < 0.05). All data shows mean ± SEM. Dpi = days post inoculation.

Figure 4

TSE vacuolation profiles in wild-type mice from the first and second mouse subpassage of brain and spleen homogenates from four cases of vCJD. The four cases comprise: an asymptomatic MV blood recipient (MV^R), the MM donor to that individual (MM^D), an MM blood donor (MM^D2008) and the associated MM blood recipient (MM^R2008). (A–C) First mouse subpassage from RIII mice. (D–F) Second mouse subpassage from RIII mice. (G–I) First mouse subpassage from C57BL6 mice. (J–L) Second mouse subpassage from C57BL6 mice. (M–O) First mouse subpassage from VM mice. (P–R) Second mouse subpassage from VM mice. Arrows indicates changes from typical vacuolation profiles. All vacuolation profiles were calculated from n ≥ 6, data show mean ± SEM. Brain region areas: G1–9, grey matter scoring areas; G1, medulla; G2, cerebellum; G3, superior colliculus; G4, hypothalamus; G5, thalamus; G6, hippocampus; G7, septum; G8, retrosplenial and adjacent motor cortex; G9, cingulate and adjacent motor cortex. W1–W3, white matter scoring regions: W1, cerebellar white matter; W2, mesencephalic tegmentum; W3, cerebral peduncle.

Figure 5

Abnormal PrP deposition in wild-type mice from the first and second mouse subpassage of brain and spleen homogenate from three cases of vCJD. The three cases comprise: an asymptomatic MV blood recipient (MV^R), the MM donor to that individual (MM^D CNS) and an MM blood donor (MM^D2008). (A) RIII mice from the first and (B) second subpassage of MVR. (C) VM mice from the first and (D) second subpassage of MV^R (E) RIII mice from the first and (F) second subpassage of MM^D (CNS). (G) VM mice from the first and (H) second subpassage of MM^D (CNS). (I) RIII mice from the first and (J) second subpassage of MM^D2008. (K) VM mice from the first and (L) second subpassage of MM^D20008 Scale bars = 200 µm (A–I).

Figure 6

Western blot analysis of brain extracts from the second mouse subpassage of RIII (A) and VM (B) wild-type mice challenged with three cases of vCJD. The three cases comprised: an asymptomatic MV blood recipient (MV^R), the MM donor to that individual [MM^D (CNS) and an MM blood donor (MM^D2008)]. A human vCJD standard positive control is shown for reference with the typical abnormal prion protein (PrP^res) type 2B. All samples were treated with proteinase K (indicated by plus symbol). The anti-prion protein detection antibody used was 6H4. Molecular weight markers are shown in kDa. PK = proteinase K.