Combinational IL-2/IL-15 induction does not further enhance IL-15-induced lymphokine-activated killer cell cytotoxicity against human leukemia/lymphoma cells

In vitro comparisons of induction of perforin (PFP), granzyme B (GRB), production of cytokines, and cell-mediated cytotoxicity by interleukin-2 (IL-2), interleukin-15 (IL-15), or combinational IL-2/IL-15-induced lymphokine-activated killer cells were studied in this study. Whereas IL-2-induction was associated with a decrease in cultured cell population over a 7-day period, IL-15 alone or in combination with IL-2 resulted in significant increase including cytotoxic T lymphocytes and subsets of CD56+ lymphocytes, particularly cytokine-induced killer and cytolytic natural killer-T lymphocytes. The overall PFP, GRB, and tumor necrosis factor-alpha expression in different subtypes were also significantly higher with IL-15 alone or in combination with IL-2 induction with resultant superior cytotoxicity compared to IL-2 treatment. There was no significant advantage of addition of IL-2 over IL-15 induction. These results offer further information on the cytotoxic potency of these cytokines and their mechanisms of action implicating potential use of IL-15 as part of cytokine adoptive immunotherapy.     

Characterisation of non-multiresistant methicillin-resistant Staphylococcus aureus (including EMRSA-15) in Kuwait Hospitals

This study characterised non-multiresistant methicillin-resistant Staphylococcus aureus (nmMRSA) isolates from Kuwait hospitals to ascertain whether they were community-acquired MRSA (CA-MRSA). Forty-two nmMRSA isolates obtained between July 2001 and October 2003 were analysed by staphylococcal cassette chromosome mec (SCCmec) typing, bacteriophage typing, production of Panton-Valentine leukocidin (PVL), urease and staphylococcal enterotoxins A, B, C and D, TSST-1, and by pulsed-field gel electrophoresis (PFGE). Forty-one isolates were SCCmec type IV, and one isolate was SCCmec type III. The isolates belonged to six PFGE patterns, with two types, A and D, distributed in six and four hospitals, respectively. Most (n = 26; 61.9%) isolates produced urease. These isolates were mainly from wound and skin infections, showed low-level methicillin resistance (MIC 8-48 mg/L), and nine carried genes for PVL. These characteristics, together with their carriage of the type-IV SCCmec, identified the isolates as CA-MRSA. Ten of the 16 urease-negative isolates produced staphylococal enterotoxin C; 12 reacted weakly with phage 75, and were resistant to clindamycin and/or erythromycin, which are characteristics of EMRSA-15. Thus, this study identified the co-existence of two types of nmMRSA, i.e., CA-MRSA and EMRSA-15, in Kuwait hospitals.     

A case of Prader – Willi syndrome arising as a result of familial unbalanced translocation t(11;15)(q25;q13)

We report on a case of Prader-Willi syndrome (PWS) with a true reciprocal unbalanced translocation, 45,XX,-15,der(11)t(11;15)pat. The proposita was diagnosed clinically as having severe PWS. Molecular studies revealed loss of the paternal methylation pattern at locus D15S63 and a deletion encompassing the loci from at least D15S10 to D15S97 of paternal chromosome 15. FISH studies confirmed the deletion of 15q11-q13 region and the presence of two telomeres on all chromosomes. The proposita's father, the father's sister and their mother are all carriers of the same balanced translocation t(11;15)(q25;q13). By genomic imprinting we would expect that if the father's sister were to give birth to a child with the same unbalanced translocation as the proband, it would be affected by Angelman syndrome.
To date, a similar familial unbalanced translocation due to loss of the small chromosome 15 derivative has not been described.     

Segregation of a supernumerary del(15) marker chromosome in sperm

Supernumerary marker chromosomes (SMC) can be associated with both normal and abnormal phenotypes. In addition, SMC are found at higher frequency in males with infertility. We identified a SMC, characterized as a del(15)(q11.2) chromosome, in a phenotypically normal male. Using fluorescence in situ hybridization (FISH), we examined the segregation of the del(15) chromosome in sperm from this patient. Only 6.23% of sperm nuclei showed disomy using a chromosome 15 alpha-satellite FISH probe, instead of the expected 50%. In addition, FISH analysis showed no increase for non-disjunction of chromosome 18, excluding an interchromosomal effect for this chromosome. The significant decrease in sperm bearing the del(15) may be due to tissue-specific mosaicism or a result of some form of selection against the del(15) during spermatogenesis. This finding provides a basis for the observation that SMC(15) are less likely to be inherited from a paternal carrier.     

Primary sequence and functional expression of a novel β subunit of the P-ATPase gene family

The cortical collecting tubule (CCT) of the mammalian kidney reabsorbs sodium and potassium, processes that are mediated by Na/K-ATPase and H/K-ATPase. CCT is also an important site for proton secretion, which is driven, in part, by H/K-ATPase. Na/K-ATPase and H/K-ATPase are members of the ion-motive P-ATPase gene family. They are closely related plasma membrane proteins which consist of heterodimers. The urinary bladder of the toad Bufo marinus is the amphibian counterpart of mammalian CCT. We have previously characterized a ouabain-resistant Na/K-ATPase [see ref. 17], from TBM cells, a clonal cell line derived from the toad bladder, which expresses transepithelial sodium transport. In the present study, we report the primary sequence and functional expression of a novel subunit ( _bladder= _bl) isolated from a toad bladder epithelial cell cDNA library. The deduced polypeptide is 299 amino acids in length and has a predicted molecular mass of 33 kDa. The _bl protein exhibits 35% amino acid identity to the previously characterized ₁ of B. marinus Na/K-ATPase and 39% identity with ₃ of B. marinus Na/K-ATPase. It shares 38% identity with the mammalian gastric H/K-ATPase and 52% with the mammalian ₂ Na/K-ATPase. Northern blot analysis shows that a 1.4×10³-base mRNA is expressed at a high level in bladder epithelial cells and eye and at a trace level in kidney; it is not detectable in significant amounts in the stomach, colon and small intestine. The _bl subunit can associate with the ₁ subunit of B. marinus Na/K-ATPase to form a functional sodium pump in the Xenopus laevis oocyte. Our data indicate that, in addition to the known ₁ and ₃ isoforms, a third distinct isoform of the subunit is present in the bladder epithelium. This new isoform could be functionally associated with subunits of either Na/K- or H/K-ATPase.     

Mammary foam cells

 Cells showing abundant, finely vacuolized cytoplasm (foam cells) are found frequently in most benign lesions of the breast and in certain malignant breast tumours. The origin of mammary foam cells (FCs) has not been clarified, and we therefore studied the morphological features of mammary FCs in a series of 50 benign lesions. The FCs were subdivided, on the basis of their distribution into FCs lining the glandular lumina, intraluminal FCs, intraepithelial-pagetoid FCs, and stromal FCs. The lesions were tested with a panel of antibodies against macrophage (MAC 387, CD68) and epithelial (epithelial membrane antigen [EMA], gross cystic disease fluid protein 15 [GCDFP15] and cytokeratin) markers. The lesions were examined for the presence of PIP/GCDFP15-specific mRNA by an in situ hybridization technique. Three different types of FCs were identified. Type A FCs are epithelial cells (positivity with EMA and cytokeratin) and show apocrine differentiation (positivity with GCDFP15 antiserum and expression of PIP/GCDFP15 mRNA). Type B FCs are of macrophage origin, as they are positive with the macrophage markers and lack cytokeratin and PIP/GCDFP15 mRNA. Finally, type C FCs show an intermediate profile between an epithelial cell and a macrophage: they are both CD68 and GCDFP15 positive and show a thin peripheral rim of positivity with anti-cytokeratin antibody. They lack PIP/GCDFP15 mRNA. Our results indicate the possibility of a spectrum of phenotypes in mammary FCs, from epithelial-apocrine cells to macrophage-derived phagocytic cells. Received: 19 September 1997 / Accepted: 29 December 1997     

DFNA54, a third locus for low-frequency hearing loss

Nonsyndromic hereditary hearing impairment (NSHHI) is a highly heterogeneous disorder with more than 90 loci mapped, of which nearly one-half of the responsible genes are identified. In dominant NSSHI hearing loss is typically biased towards the high frequencies while low-frequency hearing loss is unusual. Only two NSHHI loci, DFNA1 and DFNA6/14/38, are associated with predominantly low- frequency loss. We mapped the loci harboring the gene responsible for autosomal dominant low-frequency hearing loss in a multigenerational family. The pedigree of a Swiss family with low-frequency hearing loss was established. Using genomic DNA, DFNA1 and DFNA6/14/38 were excluded by linkage analysis or by direct sequencing of the responsible gene. Genome-wide linkage analysis was performed using commercially available microsatellite markers. Two-point linkage analysis demonstrated linkage to chromosome 5q31, the locus for DFNA15, with a lod score of 6.32 at recombination fraction =0 for marker D5S436. Critical recombinations were seen at markers D5S1972 and D5S410. Sequencing of the corresponding gene POU4F3 yielded no pathogenic mutation segregating with the affected members. In addition to Wolfram syndrome gene 1 (DFNA6/14/38) and diaphanous (DFNA1) there is evidence for a third gene involved in low-frequency hearing loss located at DFNA15. Because of the differences in auditory phenotype and the absence of pathogenic mutation in the coding region of POU4F3 it is likely that there is a second gene in 5q31, designated DFNA54, associated with NSHHI.     

IL-15真核表达质粒的构建及对乙肝疫苗诱导的体液免疫应答的影响

目的 研究两种不同的IL-15真核表达质粒对乙肝蛋白疫苗诱导的免疫应答的影响。方法：构建IL-15真核表达质粒(简称pIL-15)和含有IL-12信号肽的IL-15真核表达质粒(简称pIL-2s-15)，CTLL-2细胞增殖实验验证两种质粒真核表达产物的生物学活性。将这两种质粒分别与HBsAg共免疫BALB／C小鼠，用ELISA法检测小鼠血清抗-HBs IgG及IgGl、IgG2a亚类的效价。结果：与HBsAg蛋白疫苗共免疫时，pIL-15可使HBsAg诱导的抗-HBsIgG效价升高，显著高于载体pcDNA3．1与HB—sAg共免疫对照组，pIL-2s-15对HBsAg诱导抗-HBsIgC效价没有明显影响。与HBsAg pcDNA3．1组相比，HBsAg pIL-2s-15组和HBsAg pIL-15组诱生的抗HBsIgG2a亚类均升高，但前者IgG2a／IgG1比值最高，与HBsAg pcDNA3．1组相比差别有显著性；HBsAg pIL-15组IgG2a／IgG1比值与HBsAg pcDNA3．1组相比差别无显著性。结论 pIL-15真核表达质粒可增强蛋白疫苗诱导的体液免疫应答，pIL-2s-15真核表达质粒则主要使免疫应答趋向Th1型。     

Human CD8+ intraepithelial lymphocytes: a unique model to study the regulation of effector cytotoxic T lymphocytes in tissue

Summary:  The epithelium of the human small intestine contains a large population of intraepithelial cytolytic αβ T-cell receptor (TCR) CD8αβ T lymphocytes (IE-CTLs), whose main role is to sustain epithelial integrity by rapidly eliminating infected and damaged cells. In mouse, the recognition of inducible/modified self-molecules, i.e. non-classical major histocompatibility complex (MHC) class I molecules, is mediated by the TCR and natural killer receptors (NKRs) co-expressed on the cell surface of a non-conventional autoreactive CD8αααβTCR cell subset. In contrast, in humans, the recognition of non-classical MHC class I molecules induced by stress and inflammation on intestinal epithelial cells (IECs) is principally mediated by NKRs expressed on conventional CD8αβαβTCR cells. By sensing microenvironmental signals of inflammation and stress through NKRs, IE-CTLs fine tune their TCR activation threshold. Furthermore, IE-CTLs under particular conditions, involving interleukin-15 upregulation, acquire the capacity to kill distressed intestinal epithelial cells in an antigen non-specific manner. Adaptive IE-CTLs appear hence to have autoreactive properties and modulate their immune response based on innate signals, reflecting the fitness of the tissue.