Deletion of Stat3 enhances myeloid cell expansion and increases the severity of myeloproliferative neoplasms in Jak2V617F knock-in mice.

The JAK2V617F mutation commonly found in myeloproliferative neoplasms (MPNs) induces constitutive phosphorylation/activation of the signal transducer and activator of transcription 3 (Stat3). However, the contribution of Stat3 in MPN evoked by JAK2V617F remains unknown. To determine the role of Stat3 in JAK2V617F-induced MPN, we generated Stat3-deficient Jak2V617F-expressing mice. Whereas expression of Jak2V617F resulted in a polycythemia vera-like disease characterized by increased red blood cells (RBCs), hematocrit, neutrophils and platelets in the peripheral blood of Jak2V617F knock-in mice, deletion of Stat3 slightly reduced RBC and hematocrit parameters and modestly increased platelet numbers in Jak2V617F knock-in mice. Moreover, deletion of Stat3 significantly increased the neutrophil counts/percentages and markedly reduced the survival of mice expressing Jak2V617F. These phenotypic manifestations were reproduced upon bone marrow (BM) transplantation into wild-type animals. Flow cytometric analysis showed increased hematopoietic stem cell and granulocyte-macrophage progenitor populations in the BM and spleens of Stat3-deficient Jak2V617F mice. Stat3 deficiency also caused a marked expansion of Gr-1+/Mac-1+ myeloid cells in Jak2V617F knock-in mice. Histopathologic analysis revealed marked increase in granulocytes in the BM, spleens and livers of Stat3-deficient Jak2V617F-expressing mice. Together, these results suggest that deletion of Stat3 increases the severity of MPN induced by Jak2V617F.

Loss of wild-type Jak2 allele enhances myeloid cell expansion and accelerates myelofibrosis in Jak2V617F knock-in mice.

JAK2V617F is the most common mutation found in Philadelphia chromosome-negative myeloproliferative neoplasms (MPNs). Although a majority of MPN patients carry heterozygous JAK2V617F mutation, loss of heterozygosity (LOH) on chromosome 9p (9pLOH) involving the JAK2 locus has been observed in ∼30% of MPN patients. JAK2V617F homozygosity via 9pLOH has been associated with more severe MPN phenotype. However, the contribution of 9pLOH in the pathogenesis of MPNs remains unclear. To investigate the roles of wild-type JAK2 (JAK2 WT) and JAK2V617F alleles in the development of MPNs, we have used conditional Jak2 knock-out and Jak2V617F knock-in mice and generated heterozygous, hemizygous and homozygous Jak2V617F mice. Whereas heterozygous Jak2V617F expression results in a polycythemia vera-like MPN in mice, loss of Jak2 WT allele in hemizygous or homozygous Jak2V617F mice results in markedly increased white blood cells, neutrophils, reticulocytes and platelets in the peripheral blood, and significantly larger spleen size compared with heterozygous Jak2V617F mice. Hemizygous or homozygous Jak2V617F mice also exhibit accelerated myelofibrosis compared with mice expressing heterozygous Jak2V617F. Together, these results suggest that loss of Jak2 WT allele increases the severity of the MPN. Thus, the Jak2 WT allele functions as a negative regulator of MPN induced by Jak2V617F.

Enterotoxin-specific immunoglobulin E responses in humans after infection or vaccination with diarrhea-causing enteropathogens.

Cholera toxin (CT)-specific antibody responses of the immunoglobulin E (IgE) isotype in the sera of adult patients suffering from infection with either Vibrio cholerae O1, V. cholerae O139, or enterotoxigenic Escherichia coli (ETEC) were analyzed and compared with those in the sera of volunteers immunized with a bivalent B subunit O1/O139 whole-cell cholera vaccine. A significant IgE response to CT was observed in 90% of the patients with V. cholerae O1 infection (18 of 20; P = <0.001) and 95% of the patients with V. cholerae O139 infection (19 of 20; P = <0.001). Similarly, the majority of the patients with ETEC diarrhea (83%; 13 of 15) showed a positive IgE response to CT. Eight of 10 North American volunteers (80%) orally challenged with V. cholerae O1 showed CT-specific IgE responses (P = 0.004). In contrast, Swedish volunteers immunized with the oral cholera vaccine showed no IgE responses to CT (P value not significant). During the study period, total IgE levels in the sera of the diarrheal patients, the North American volunteers, and the Swedish cholera vaccinees alike remained unchanged. However, the total IgE levels in the sera of patients and healthy Bangladeshi controls were on average 89-fold higher than those in the sera of the healthy Swedish volunteers and 34-fold higher than those in the sera of the North American volunteers.

Comparison of immune responses in patients infected with Vibrio cholerae O139 and O1.

Vibrio cholerae O139 has recently emerged as the second etiologic agent of cholera in Asia. A study was carried out to evaluate the induction of specific immune responses to the organism in V. cholerae O139-infected patients. The immune responses to V. cholerae O139 Bengal were studied in patients by measuring antibody-secreting cells (ASC), as well as vibriocidal and antitoxic antibodies in the circulation. These responses were compared with those in patients with V. cholerae O1 disease. Strong immunoglobulin A (IgA) and IgM ASC responses were seen against the homologous lipopolysaccharide or serogroup of V. cholerae. The magnitude and isotype of the responses were similar in O139- and O1-infected patients. Vibriocidal antibody responses were seen against bacteria of the homologous but not heterologous serogroup, and these responses reflect the lack of cross-protection between the infections caused by the two serogroups. The two groups of patients showed comparable cholera toxin-specific ASC responses, with the IgG isotype dominating over the IgA isotype, as well as comparable antitoxic immune responses in plasma. These results suggest that despite having a polysaccharide capsule, V. cholerae O139 induces systemic and intestine-derived ASC responses in peripheral blood comparable to those seen in patients with V. cholerae O1 disease.

Comparison of the vibriocidal antibody response in cholera due to Vibrio cholerae O139 Bengal with the response in cholera due to Vibrio cholerae O1.

Vibrio cholerae serogroup O139, now considered to be the second organism capable of causing epidemic severe dehydrating cholera, contains a capsular polysaccharide which makes it difficult for it to be used in the conventional vibriocidal antibody assay optimized for V. cholerae O1. After modification of the procedure, which involved the use of specific bacterial strains, a lower bacterial inoculum, and increased amounts of complement, the vibriocidal antibody responses to V. cholerae O139 were measured in acute- and convalescent-phase sera from 33 V. cholerae O139-infected and 18 V. cholerae O1-infected patients and in single serum samples from 20 healthy control subjects. The responses in these individuals to V. cholerae O1 strains were also determined. Significant elevations in the homologous antibody response were found only in the convalescent-phase sera from both groups of patients with cholera. These findings may explain the basis for the lack of heterologous protection between the two serogroups of V. cholerae. Healthy controls had higher background levels of vibriocidal antibody to V. cholerae O1 than to V. cholerae O139.