Loss of B Cells in Patients with Heterozygous Mutations in IKAROS.

BACKGROUND: Common variable immunodeficiency (CVID) is characterized by late-onset hypogammaglobulinemia in the absence of predisposing factors. The genetic cause is unknown in the majority of cases, and less than 10% of patients have a family history of the disease. Most patients have normal numbers of B cells but lack plasma cells. METHODS: We used whole-exome sequencing and array-based comparative genomic hybridization to evaluate a subset of patients with CVID and low B-cell numbers. Mutant proteins were analyzed for DNA binding with the use of an electrophoretic mobility-shift assay (EMSA) and confocal microscopy. Flow cytometry was used to analyze peripheral-blood lymphocytes and bone marrow aspirates. RESULTS: Six different heterozygous mutations in IKZF1, the gene encoding the transcription factor IKAROS, were identified in 29 persons from six families. In two families, the mutation was a de novo event in the proband. All the mutations, four amino acid substitutions, an intragenic deletion, and a 4.7-Mb multigene deletion involved the DNA-binding domain of IKAROS. The proteins bearing missense mutations failed to bind target DNA sequences on EMSA and confocal microscopy; however, they did not inhibit the binding of wild-type IKAROS. Studies in family members showed progressive loss of B cells and serum immunoglobulins. Bone marrow aspirates in two patients had markedly decreased early B-cell precursors, but plasma cells were present. Acute lymphoblastic leukemia developed in 2 of the 29 patients. CONCLUSIONS: Heterozygous mutations in the transcription factor IKAROS caused an autosomal dominant form of CVID that is associated with a striking decrease in B-cell numbers. (Funded by the National Institutes of Health and others.).

The effect of Acemannan Immunostimulant in combination with surgery and radiation therapy on spontaneous canine and feline fibrosarcomas.

Eight dogs and five cats with histopathologically confirmed fibrosarcomas were treated with Acemannan Immunostimulanta in combination with surgery and radiation therapy. These animals had recurring disease that had failed previous treatment, a poor prognosis for survival, or both. Following four to seven weekly acemannan treatments, tumor shrinkage occurred in four (greater than 50%; n = 2) of 12 animals, with tumors accessible to measurement. A notable increase in necrosis and inflammation was observed. Complete surgical excision was performed on all animals between the fourth and seventh week following initiation of acemannan therapy. Radiation therapy was instituted immediately after surgery. Acemannan treatments were continued monthly for one year. Seven of the 13 animals remain alive and tumor-free (range, 440+ to 603+ days) with a median survival time of 372 days. The data suggests that Acemannan Immunostimulant may be an effective adjunct to surgery and radiation therapy in the treatment of canine and feline fibrosarcomas.

Effect of incorporation of serum from dogs with renal impairment on canine erythroid bone marrow cultures.

Serum from dogs with surgically induced renal impairment was incorporated into the medium for erythroid bone marrow cultures. A significant correlation was found between serum activities of erythropoietin and numbers of erythroid colony-forming units grown in culture. Serum creatinine concentrations had no correlation, and serum parathyroid hormone activities had a negative correlation with numbers of erythroid colony-forming units that was below the level of significance. Purified 1-84 parathyroid hormone added to bone marrow cultures was found to be stimulatory to erythroid colony-forming unit growth in higher concentrations, but decreased the number of burst-forming units. Unmeasured substances in the canine serum appeared to have a greater effect on the canine erythroid bone marrow cultures than did creatinine or parathyroid hormone values.

Effects of estradiol on hematopoietic and marrow adherent cells of dogs.

Hematopoietic and marrow adherent cells were examined in 5 ovariohysterectomized dogs treated once with 1.5 mg of 17 beta-estradiol cyclopentylpropionate/kg of body weight. All dogs initially developed thrombocytopenia and neutrophilic leukocytosis, with megakaryocytopenia and an increased myeloid/erythroid ratio of the marrow. Later, 2 dogs became thrombocytopenic and neutropenic, with megakaryocytopenia, decreased myeloid/erythroid ratio, and increased concentration of plasma cells and mast cells in the marrow. The concentration of marrow granulocyte-macrophage colony-forming units (CFU-GM) was decreased at 2 and 3 weeks in all dogs after estradiol treatment. The concentration of marrow fibroblast CFU-F was decreased in all estradiol-treated dogs at 1 and 2 weeks. The ability of marrow adherent cells to support CFU-GM colony formation was unchanged in estradiol-treated dogs. Estradiol sulfate at final concentrations of 10(-8) to 10(-4)M did not affect the in vitro growth of CFU-GM and CFU-F, or the hematopoietic ability of marrow adherent cells from nontreated dogs.

Myelopoiesis and marrow adherent cells in estradiol-treated mice.

Myelopoiesis and marrow adherent cells were evaluated in C57Bl/6J mice at two and four weeks after treatment with 0.1 mg 17 beta-estradiol cyclopentylpropionate. Estradiol-treated mice were lymphopenic and eosinopenic at two and four weeks; in addition, neutropenia occurred at four weeks. Numbers of lymphoid, granulocytic, and erythroid cells were decreased in the marrow at two and four weeks. The numbers of granulocyte-macrophage and fibroblast colony-forming units in the humeral marrow were also decreased at two and four weeks. However, the hematopoietic ability of marrow adherent cells was unchanged in estradiol-treated mice. Thymic cortical atrophy, metaphyseal osteosclerosis, and neutrophilic infiltration of the uterus occurred in estradiol-treated mice.

Response to neonatally thymectomized Holstein calves to bacille Calmette Guérin.

Neonatally thymectomized and intact Holstein calves were inoculated intradermally with bacille Calmette Guérin mycobacteria at 4 to 6 months of age. The immune response of the calves was evaluated by in vitro lymphocyte stimulation, with a purified protein derivative of Mycobacterium bovis, and by a cervical skin test. Eight of te calves had been previously infected at 7 to 15 weeks of age with bovine viral diarrhea virus. The cutaneous and in vitro immune responses were not different in neonatally thymectomized calves, whether or not they had been previously infected with bovine viral diarrhea virus. However, a prominent histologic difference in the nature of the primary granulomatous lesions was observed in the thymectomized calves. These injection sites were characterized by few lymphocytes and an increased number of multinucleated giant cells. These observations were interpreted as manifestations of a deficient thymus-dependent effector response in the thymectomized calves.

Increased incidence of bovine papular stomatitis in neonatal calves.

A high incidence of bovine papular stomatitis (BPS) occurred in neonatal calves following neonatal thymectomy and antilymphocyte globulin treatment, sham thymectomy, treatment with normal horse globulin and in untreated calves. The source of BPS virus was not identified but was suspected to be latent in the calves and activated by thymectomy although no experimental evidence directly supported this conclusion. The potential for activation of a latent BPS infection was indicated by an apparent relationship between the stress of surgery or foreign protein inoculation and the severity of lesions. Subsequent bovine viral diarrhea virus infection did not result in recrudescence.

Acute experimental canine ehrlichiosis. II. Sequential reaction of the hemic and lymphoreticular system of selectively immunosuppressed dogs.

The lymphoreticular response of dogs to Ehrlichia canis infection was studied after immunosuppressive therapy with cyclophosphamide or antilymphocyte serum. Immunosuppression did not increase mortality, nor did it prevent or significantly modify the clinical manifestations of acute ehrlichiosis. The clinical, hematologic and serum biochemical characteristics of acute ehrlichiosis appeared seven or more days later in immunosuppressed than in immunologically intact infected dogs. Immunosuppressive therapy generally attenuated the lesions of acute ehrlichiosis. Lymphoreticular tissues were partially depleted of lymphocytes, as would be expected following cyclophosphamide therapy, but the hemopoietic tissue of cyclophosphamide-treated principals was much more cellular than that of the controls, perhaps demonstrating a direct stimulatory effect E. canis infection on bone marrow. Antilymphocyte serum therapy reduced cellularity of the thymic-dependent areas in both the spleen and lymph nodes, but B-cell areas were not affected. The B-cell areas did not increase in activity during infection in the antilymphocyte serum-treated dogs, suggesting the need for an intact T-cell compartment for the lymphocytic proliferation that occurs in nonsuppressed infected dogs. Renal vasculitis and lympho-plasmacytosis were absent and the pulmonary and hepatic lesions were not as pronounced in the dogs given antilymphocyte serum as in immunologically competent infected dogs.

Serum immunoglobulin concentrations of cattle in a herd with bovine leukosis.

Consistent relationship was not found between the serum concentrations of immunoglobulin IgM and IgG and the concentration of blood lymphocytes (Bendixen index) in 289 Holstein-Friesian heifers and cows from a dairy herd in which bovine lymphoma had been identified repeatedly. Serum IgM was present in all cattle, and those cows with highest concentrations of blood lymphocytes generally had higher levels of IgM.