Murine JAK3 is preferentially expressed in hematopoietic tissues and lymphocyte precursor cells

To elucidate the role of cytokine receptor signal transduction in T- cell development, we have investigated the expression pattern and biochemical characteristics of the murine Janus family tyrosine kinase, JAK3. Previous studies have shown that JAK3 is expressed in lymphoid and myeloid tumor cell lines and in a small number of lymphoid tissues. To further characterize JAK3 expression, we used a quantitative polymerase chain reaction approach to compare JAK3 mRNA levels at multiple stages of T-cell differentiation and in a broad range of mouse tissues. These studies, in conjunction with analyses of JAK3 protein expression, show that the highest levels of JAK3 are in adult, 2-week- old, and fetal thymus, followed by somewhat lower levels in bone marrow, spleen, fetal liver, and adult CD4-CD8- thymocytes. We also show that different forms of JAK3 mRNA arise by alternative splicing. Finally, our biochemical studies show that the JAK3 kinase domain, but not the pseudo-kinase domain, has tyrosine kinase activity and, furthermore, that JAK3 kinase activity is abolished by an amino acid substitution of the conserved lysine in the kinase domain (K851R). These studies show that JAK3 expression is profoundly skewed to hematopoietic and lymphoid precursor cells, strongly suggesting a role for JAK3 in hematopoiesis and T- and B-cell development.     

Hepatosplenic T-cell lymphoma: a distinct clinicopathologic entity of cytotoxic gamma delta T-cell origin

We identified eight cases of T-cell lymphoma with evidence of a gamma delta phenotype over a 13-year period. Seven of these cases conformed to a distinct clinicopathologic entity of hepatosplenic gamma delta T- cell lymphoma. Nearly all of these patients were young adult males (five of seven), with a median age at presentation of 20 years. They presented with marked hepatosplenomegaly, without lymphadenopathy or significant peripheral blood lymphocytosis. Thrombocytopenia was seen in all patients, and five of seven were mildly anemic. The clinical course was aggressive, and despite multiagent chemotherapy, the median survival duration was less than 1 year. The morphologic findings were uniform; a monomorphic population of medium-sized lymphoid cells with moderately clumped chromatin and a rim of pale cytoplasm infiltrated the sinusoids of the spleen, liver, and bone marrow. The cells had a characteristic immunophenotype: CD2+, CD3+, CD4-, CD5-, CD7+, CD16+, CD57-, CD25-, T-cell receptor (TCR)delta +, beta F1-. CD8 was positive in four of seven cases tested, and CD56 was positive in five of six. All cases expressed the cytotoxic granule-associated protein, TIA1, but perforin was detected in only one case. All cases with assessable DNA had a TCR gamma gene rearrangement, and lacked Epstein-Barr virus sequences. Isochromosome 7q was identified in two cases with cytogenetic information. The one case of cutaneous gamma delta T-cell lymphoma differed in its clinical manifestations, histologic appearance, and immunophenotype. We conclude that hepatosplenic gamma delta T-cell lymphoma is a distinct clinicopathologic entity derived from cytotoxic gamma delta T cells, and should be distinguished from other lymphomas of T-cell and natural-killer cell (NK)-like T-cell derivation.     

Paediatric nodal marginal zone B‐cell lymphadenopathy of the neck: a Haemophilus influenzae‐driven immune disorder?

Many hyperplasias and lymphomas of marginal zone B‐cells are associated with infection. We identified six children and one adolescent with cervical lymphadenopathy showing prominent polyclonal nodal marginal zone hyperplasia (pNMZH) and four adolescents with monoclonal paediatric nodal marginal zone lymphoma (pNMZL). The clonality status was assessed using BIOMED‐2‐IG PCR analysis. Haemophilus influenzae was identified in all six cases of pNMZH that could be tested by direct culture (N = 3) or a very sensitive PCR for the H. influenzae gyrase gene in frozen materials (N = 5). H. influenzae was not detected in three pNMZLs and 28 non‐specific reactive cervical lymph nodes of age‐matched controls, except for a single control node that was obtained during oropharyngeal surgery for a cleft palate showing very low copy numbers of H. influenzae. pNMZH patients were younger than pNMZL patients (median age 12 versus 21 years). pNMZH showed a prominent nodular appearance with variable fibrosis without acute inflammation. Within the nodules, the expanded germinal centres and variably sized marginal zones were colonized by activated B‐cells with weak expression of IgD and lack of CD10 and/or BCL6 expression. Some areas showed skewed light chain expression in plasma cells (4/5 cases lambda). In four cases tested, this was confirmed by flow cytometry for surface Ig (3/4 cases lambda). In contrast, pNMZL showed more extensive expansion of marginal zones by centrocytoid cells and often expression of BCL2 protein. Several H. influenzae strains are known to interact with the constant part of IgD on human B‐cells, leading to their polyclonal proliferation and activation. We speculate that in vivo stimulation of IgD+ marginal zone B‐cells by this bacterium may be implicated in this particular lymphadenopathy that should be distinguished from monoclonal pNMZL. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Surveillance of active human cytomegalovirus infection in hematopoietic stem cell transplantation (HLA sibling identical donor): search for optimal cutoff value by real-time PCR

Background  

Human cytomegalovirus (CMV) infection still causes significant morbidity and mortality after allogeneic hematopoietic stem cell transplantation (HSCT). Therefore, it is extremely important to diagnosis and monitor active CMV infection in HSCT patients, defining the CMV DNA levels of virus replication that warrant intervention with antiviral agents in order to accurately prevent CMV disease and further related complications.     

Cyclosporin as initial treatment for Crohn's disease

Childhood Crohn's disease may cause significant morbidity. T cell activation is considered to be central to Crohn's disease pathology, and as cyclosporin is a powerful inhibitor of T cell activation, and has been used in adult Crohn's disease with encouraging results, it may offer the prospect of remission if given early in the course of disease. Children with newly diagnosed Crohn's disease or those relapsing off treatment were therefore given cyclosporin or conventional treatment (enteral nutrition or corticosteroids) by random allocation. Evaluation was performed initially and at two months. Twenty four children were studied (10 on cyclosporin and 14 on conventional treatment; one child on cyclosporin withdrew). Significant clinical improvement occurred in the group on conventional treatment, but not in the cyclosporin group. Colonoscopic improvement was noted in 5/9 on cyclosporin and 8/14 on conventional treatment, but neither group produced a significant fall in median colonoscopic index. Histological improvement was seen in 7/8 on cyclosporin and 8/13 on conventional treatment, but cyclosporin was not significantly better. Cyclosporin produced improved clinical and histological appearance without matched improvement in blood disease indices. It was not better than conventional treatment, and simple oral administration is probably not suitable for newly diagnosed patients with Crohn's disease.     

Insulin-like growth factor (IGF)-I and IGF binding protein-3 concentrations in fluid from human stimulated follicles

Insulin-like growth factor-I (IGF-I) and IGF binding protein-3 (IGFBP- 3) play an important role in regulating follicle growth and maturation. We have evaluated whether responsiveness to gonadotrophins during an in- vitro fertilization (IVF) treatment is related to follicular fluid IGF- I and IGFBP-3 concentrations. We also investigated if a difference is present in IGF-I and IGFBP-3 concentrations between patients treated with human menopausal gonadotrophin (HMG) and patients treated with highly purified follicle stimulating hormone (FSH). We have measured IGF-I and IGFBP-3 in follicular fluid from pre-ovulatory follicles in an IVF programme. All 70 patients were stimulated after being down- regulated with a gonadotrophin-releasing hormone (GnRH) analogue. IGF-I concentrations in follicular fluid were significantly inversely correlated with the number of ampoules FSH administered and number of days of FSH administration, and significantly correlated with the number of follicles aspirated. IGFBP-3 concentrations were not correlated with any other parameter measured nor were IGF-I and IGFBP-3 concentrations correlated. IGFBP-3 concentrations were significantly higher in patients receiving highly purified FSH compared with patients receiving HMG (P < 0.005). These results are new evidence that IGF-I concentration in follicular fluid is higher in women who respond better to follicular stimulation, i.e. women who grow many follicles, women who need a shorter duration of stimulation and women who need fewer ampoules FSH before oocyte retrieval.      

Altered metabolism of familial Alzheimer's disease-linked amyloid precursor protein variants in yeast artificial chromosome transgenic mice

Missense mutations in the beta-amyloid precursor protein gene (APP) co- segregate with a small subset of autosomal dominant familial Alzheimer's disease (FAD) cases wherein deposition of the 39-43 amino acid beta-amyloid (A beta) peptide and neurodegeneration are principal neuropathological hallmarks. To accurately examine the effect of missense mutations on APP metabolism and A beta production in vivo, we have introduced yeast artificial chromosomes (YACs) containing the entire approximately 400 kbp human APP gene encoding APP harboring either the asparagine for lysine and leucine for methionine FAD substitution at codons 670 and 671 (APP(K670N/M671L)), the isoleucine for valine FAD substitution at codon 717 (APP(V7171)) or a combination of both substitutions into transgenic mice. We demonstrate that, relative to YAC transgenic mice expressing wild-type APP, high levels of A beta peptides are detected in the brains of YAC transgenic mice expressing human APP(K670N/M671L) that is associated with a concomitant diminution in the levels of apha-secretase-generated soluble APP derivatives. Moreover, the levels of longer A beta peptides (species terminating at amino acids 42/43) are elevated in YAC transgenic mice expressing human APP(V7171). These mice should prove valuable for detailed analysis of the in vivo effects of the APP FAD mutations in a variety of tissues and throughout aging and for testing therapeutic agents that specifically alter APP metabolism and A beta production.      

Autosomal recessive retinitis pigmentosa and cone-rod dystrophy caused by splice site mutations in the Stargardt's disease gene ABCR

Ophthalmological and molecular genetic studies were performed in a consanguineous family with individuals showing either retinitis pigmentosa (RP) or cone-rod dystrophy (CRD). Assuming pseudodominant (recessive) inheritance of allelic defects, linkage analysis positioned the causal gene at 1p21-p13 (lod score 4.22), a genomic segment known to harbor the ABCR gene involved in Stargardt's disease (STGD) and age- related macular degeneration (AMD). We completed the exon-intron structure of the ABCR gene and detected a severe homozygous 5[prime] splice site mutation, IVS30+1G->T, in the four RP patients. The five CRD patients in this family are compound heterozygotes for the IVS30+1G- >T mutation and a 5[prime] splice site mutation in intron 40 (IVS40+5G- >A). Both splice site mutations were found heterozygously in two unrelated STGD patients, but not in 100 control individuals. In these patients the second mutation was either a missense mutation or unknown. Since thus far no STGD patients have been reported to carry two ABCR null alleles and taking into account that the RP phenotype is more severe than the STGD phenotype, we hypothesize that the intron 30 splice site mutation represents a true null allele. Since the intron 30 mutation is found heterozygously in the CRD patients, the IVS40+5G->A mutation probably renders the exon 40 5[prime] splice site partially functional. These results show that mutations in the ABCR gene not only result in STGD and AMD, but can also cause autosomal recessive RP and CRD. Since the heterozygote frequency for ABCR mutations is estimated at 0.02, mutations in ABCR might be an important cause of autosomal recessive and sporadic forms of RP and CRD.