Protective heterologous antiviral immunity and enhanced immunopathogenesis mediated by memory T cell populations

A basic principle of immunology is that prior immunity results in complete protection against a homologous agent. In this study, we show that memory T cells specific to unrelated viruses may alter the host's primary immune response to a second virus. Studies with a panel of heterologous viruses, including lymphocytic choriomeningitis (LCMV), Pichinde (PV), vaccinia (VV), and murine cytomegalo (MCMV) viruses showed that prior immunity with one of these viruses in many cases enhanced clearance of a second unrelated virus early in infection. Such protective immunity was common, but it depended on the virus sequence and was not necessarily reciprocal. Cell transfer studies showed that both CD4 and CD8 T cell populations from LCMV-immune mice were required to transfer protective immunity to naive hosts challenged with PV or VV. In the case of LCMV-immune versus naive mice challenged with VV, there was an enhanced early recruitment of memory phenotype interferon (IFN) gamma-secreting CD4(+) and CD8(+) cells into the peritoneal cavity and increased IFN-gamma levels in this initial site of virus replication. Studies with IFN-gamma receptor knockout mice confirmed a role for IFN-gamma in mediating the protective effect by LCMV-immune T cell populations when mice were challenged with VV but not PV. In some virus sequences memory cell populations, although clearing the challenge virus more rapidly, elicited enhanced IFN-gamma-dependent immunopathogenesis in the form of acute fatty necrosis. These results indicate that how a host responds to an infectious agent is a function of its history of previous infections and their influence on the memory T cell pool.     

Progesterone induces calcitonin gene expression in human endometrium within the putative window of implantation

The human endometrium acquires the ability to implant the developing embryo within a specific time window that is thought to open between days 19-24 of the secretory phase of the menstrual cycle. During this period the endometrium undergoes pronounced structural and functional changes induced by the ovarian steroids, estrogen and progesterone, that prepare it to be receptive to invasion by the embryo. The identification of reliable biochemical markers to assess this critical receptive phase in the context of the natural cycle remains one of the major challenges in the study of human reproduction. Our previous studies in a rat model system demonstrated that the expression of calcitonin, a peptide hormone involved in calcium homeostasis, is transiently induced by progesterone in the glandular epithelium at the onset of implantation. Attenuation of calcitonin synthesis in the uterus during the preimplantation phase by administration of calcitonin antisense oligodeoxynucleotides severely impairs implantation of rat embryos, suggesting that this peptide hormone plays a critical role in uterine receptivity. To investigate whether calcitonin is also expressed in the human endometrium during implantation, we monitored the spatio-temporal expression of calcitonin on various days of the menstrual cycle. Our studies employing RT-PCR showed that calcitonin messenger ribonucleic acid is expressed in human endometrium during the postovulatory midsecretory phase (days 17-25) of the menstrual cycle, with maximal expression occurring between days 19-21. Very little calcitonin expression was detected in the endometrium in either the preovulatory proliferative (days 5-14) or the late secretory (days 26-28) phase. In situ hybridization and immunocytochemical analyses localized the calcitonin expression predominantly in the glandular epithelial cells of the endometrium. Our studies further showed that calcitonin expression in the human endometrium is under progesterone regulation. Treatment of women with an antiprogestin, mifepristone (RU-486), drastically reduced calcitonin expression in the endometrium. Collectively, these findings reveal that progesterone-induced expression of calcitonin in the secretory endometrium temporally coincides with the putative window of implantation in the human.     

Tie-over bolster dressings may not be necessary to secure small full thickness skin grafts

BACKGROUND: The accepted method for securing full thickness skin grafts (FTSG) is with a tie-over bolster dressing, with or without basting sutures. We question the need for tie-over bolster dressings for small FTSGs. OBJECTIVE: We describe our method of FTSG reconstruction and the outcome in a consecutive series. METHODS: Thirty patients with surgical defects following tumour excision from the face24, scalp1, ear2, and finger3 ranging in diameter from 8 to 45 mm (mean 20 mm) were reconstructed with a FTSG. Interrupted monofilament nylon perimeter sutures only were used, with antibiotic ointment at the wound edge with either a light dressing or no dressing. RESULTS: A series of 30 FTSG secured without a tie-over bolster dressing or basting sutures is described, all with good to excellent long term results. In two cases there was early superficial necrosis of the graft, but ultimately 100% graft take. CONCLUSION: We have found tie-over bolster dressings and basting sutures to be unnecessary in our series of small FTSG. This technique saves time and material and minimizes handling of the graft.     

Autophosphorylation-independent activation of Acanthamoeba myosin I heavy chain kinase by plasma membranes

The three isoforms of Acanthamoeba myosin I (non-filamentous myosin with only a single heavy chain) express actin-activated Mg(2+)-ATPase activity only when phosphorylated at a single site by myosin I heavy chain kinase. The kinase is activated by autophosphorylation that is greatly stimulated by acidic phospholipids. Substantial fractions of the three myosins I and the kinase are associated in situ with membranes, and all four enzymes bind to purified membranes in vitro. We now report that when kinase and myosin I are incubated together with phosphatidylserine vesicles not only does the kinase autophosphorylate more rapidly than soluble kinase in the absence of phosphatidylserine but that, probably as a result, the kinase phosphorylates myosin I more rapidly than soluble kinase phosphorylates soluble myosin I. Similarly, plasma membrane-bound kinase phosphorylates membrane-bound myosin I and activates its actin-activated Mg(2+)-ATPase activity more rapidly than soluble kinase phosphorylates and activates soluble myosin I in the absence of membranes. However, the enhanced activity of membrane-bound kinase (which is comparable to the activity of kinase in the presence of phosphatidylserine) is not due to autophosphorylation of the membrane-bound kinase, which is very much slower than for kinase activated by phosphatidylserine vesicles.     

Omission of temporary diversion in restorative proctocolectomy--is it safe?

PURPOSE: The aim of our study was to evaluate the safety and functional outcome of restorative proctocolectomy (RP) without diversion. METHODS: Fifty patients underwent RP without diversion for ulcerative colitis (82 percent), familial adenomatous polyposis (12 percent), and indeterminate colitis (6 percent). The perioperative course and functional outcome of these patients were compared with another group of 50 patients undergoing RP with diverting ileostomy during the same time period (1989-1991) and closely matched for age, gender, surgeon, diagnosis, extent and duration (median, 10 years) of colitis, prior colectomy (approximately 22 percent), steroid use (40 percent), type of pouch, distance of ileal pouch-anal anastomosis from the dentate line (median, 1.5 cm), and the duration of follow-up (median, 12 months). All patients had a stapled ileal pouch-anal anastomosis without mucosectomy and a smooth conduct of the operation. RESULTS: There was no operative mortality. Anastomotic leaks and pelvic abscess were more common in patients without ileostomy (7/50 or 14 percent vs. 2/50 or 4 percent); 8 of these 9 patients were taking > or = 20 mg of prednisone/day. Septic complications requiring relaparotomy (6 percent vs. 0 percent), prolonged ileus, and fever of unknown origin (10 percent vs. 4 percent) were also more common in patients without ileostomy. Despite similar functional results at 6 weeks and at 12 months after initial pouch function, patients without ileostomy had a poorer quality of life index (5 vs. 8; 10 being best) in the early period (0-6 weeks) of pouch function. CONCLUSION: In equally favorable cases, RP without diversion is not as safe as RP with diversion, especially in patients taking > or = 20 mg of prednisone/day.     

Segmental vascular pressures in lung embolism

Average microvascular filtration pressure and vascular permeability measures were obtained in 100-microns glass bead-embolized dog lung lobes randomly assigned to groups in which isolated perfusion was designed to produce weight gain (edema groups) or no weight gain (isogravimetric groups). The solvent drag reflection coefficient (sigma), an index of vascular permeability, was obtained during edema formation, whereas isogravimetric capillary pressure was obtained during isogravimetry. Vascular permeability increased in response to embolism, because sigma was 0.53 +/- 0.03 vs. 0.80 +/- 0.05 (P < 0.005) in embolized and control lobes, respectively. Vascular occlusion methods indicated the greatest resistance increase in response to embolism in the vascular segment represented by Pao--Pdo (arterial occlusion pressure--double occlusion pressure). Because papaverine vasodilation reduced total vascular resistance (RT; P < 0.05) by decreasing Pao (P < 0.01) without altering Pdo, the RT increase in response to embolism was likely due to both vasoconstriction and obstruction. Because Pdo approximated capillary pressure at isogravimetry, Pdo appears to estimate average filtration pressure in both embolized (n = 6) and control lungs (n = 6). Arterial pressure was 56.2 +/- 13.6 vs. 17.6 +/- 1.5 cmH2O (P < 0.01) in embolized (n = 5) and control lobes (n = 6), respectively, whereas Pdo values of 16.1 +/- 1.5 vs. 12.4 +/- 0.8 (P < 0.05) suggested relatively little increase in filtration pressure in response to embolism. If the beads obstructed 100-microns vessels, the vascular segment represented by Pao--Pdo, the major site of vasoconstriction as well as mechanical obstruction, likely includes 100-microns arteries.     

A new experimental in vitro culture medium for cultivation of Leishmania species

A new liquid culture medium prepared with chemicals that can be obtained economically and commercially was tested in in vitro cultivation of Leishmania promastigotes to obtain a large number of organisms to use in serological studies. The number of Leishmania infantum and Leishmania tropica promastigotes taken from Novy-MacNeal-Nicolle (NNN) medium reached 1 x 10(7)/ml at the end of the 8th day in our new medium, though in NNN medium the number of organisms reached only 5 x 10(6)/ml. After 10 subsequent passages, the culture medium prepared was evaluated as being quite inexpensive, simple, and successful compared with other commercially available liquid culture media.     

An investigation of the intra-generic structure of Rothia by pyrolysis mass spectrometry

Some women with benign breast disease eventually develop breast cancer. The mammary gland undergoes tissue remodelling according to hormonal influences, involving a balance between quiescence, proliferation, and mechanisms of cell death. Proliferation and/or apoptotic events could therefore be investigated to help understand the mechanisms of benign lesion formation and identify mastopathies with a poor prognosis. bcl-2 expression was analysed by immunohistochemistry in 75 benign mastopathies. Protein levels were quantitated with an image analyser in various epithelial structures on frozen sections, including adenoses, fibroadenomas, ductal epithelial hyperplasias, cysts, and apparently normal surrounding lobules and ducts. bcl-2 levels were equivalent in apparently normal lobules and ducts, as well as in cysts and ductal hyperplasias. bcl-2 staining was significantly higher in fibroadenomas, known to be of lobular origin [mean = 10.1, quantitative immunochemistry score (QIC) arbitrary units (AU), n = 19], than in normal lobules (mean = 5.1 AU, n = 43, P = 7 x 10(-5). bcl-2 levels in normal lobules and ducts varied according to the menstrual cycle, being higher during the follicular than the luteal phase (P = 1.8 x 10(-2) and P = 1.7 x 10(-2), respectively). This was further supported by a statistical link (P = 5 x 10(-3) between high levels of circulating progesterone and weak bcl-2 staining in lobules and ducts. This progesterone-dependent variation was absent in fibroadenomas. No statistical correlation was found between bcl-2 expression and circulating levels of oestradiol, and follicle-stimulating or luteotrophic hormones. Although these are only preliminary results, they suggest an influence of progesterone on bcl-2 expression which might be lost in fibroadenomas. A hypothesis is proposed concerning the potential involvement of altered regulation of the apoptotic process in the formation of such benign lesions.     

ABT-089 [2-methyl-3-(2-(S)-pyrrolidinylmethoxy)pyridine]: I. A potent and selective cholinergic channel modulator with neuroprotective properties

Lactate dehydrogenase from the malarial parasite Plasmodium falciparum has many amino acid residues that are unique compared to any other known lactate dehydrogenase. This includes residues that define the substrate and cofactor binding sites. Nevertheless, parasite lactate dehydrogenase exhibits high specificity for pyruvic acid, even more restricted than the specificity of human lactate dehydrogenases M4 and H4. Parasite lactate dehydrogenase exhibits high catalytic efficiency in the reduction of pyruvate, kcat/Km = 9.0 x 10(8) min(-1) M(-1). Parasite lactate dehydrogenase also exhibits similar cofactor specificity to the human isoforms in the oxidation of L-lactate with NAD+ and with a series of NAD+ analogs, suggesting a similar cofactor binding environment in spite of the numerous amino acid differences. Parasite lactate dehydrogenase exhibits an enhanced kcat with the analog 3-acetylpyridine adenine dinucleotide (APAD+) whereas the human isoforms exhibit a lower kcat. This differential response to APAD+ provides the kinetic basis for the enzyme-based detection of malarial parasites. A series of inhibitors structurally related to the natural product gossypol were shown to be competitive inhibitors of the binding of NADH. Slight changes in structure produced marked changes in selectivity of inhibition of lactate dehydrogenase. 7-p-Trifluoromethylbenzyl-8-deoxyhemigossylic acid inhibited parasite lactate dehydrogenase, Ki = 0.2 microM, which was 65- and 400-fold tighter binding compared to the M4 and H4 isoforms of human lactate dehydrogenase. The results suggest that the cofactor site of parasite lactate dehydrogenase may be a potential target for structure-based drug design.     

Hereditary isolated renal magnesium loss maps to chromosome 11q23

Dendritic cells (DC) expanded in the presence of GM-CSF from the bone marrow of C57BL/6 mice process Gram-negative bacteria expressing the model antigen Crl-OVA for peptide presentation on MHC class I molecules. Here we show that presentation of OVA(257-264) processed by DC co-incubated with E. coli expressing Crl-OVA, which contains the Kb-binding OVA(257-264) epitope, occurs by a cytosolic MHC-I presentation pathway. First, we demonstrate the requirement for the transporter associated with antigen processing (TAP) by showing that DC from TAP1-/- mice co-incubated with E. coli expressing Crl-OVA did not result in Kb presentation of OVA(257-264). Second, the proteasome inhibitor MG132 abrogated presentation of OVA(257-264) on Kb when C57BL/6 DC phagocytosed and processed E. coli expressing Crl-OVA. Third, inhibiting protein synthesis using cycloheximide or blocking exocytosis of newly synthesized proteins from the endoplasmic reticulum using brefeldin A abrogated presentation of OVA(257-264) processed from bacteria expressing Crl-OVA by C57BL/6 DC. Finally, peptide regurgitation and loading of OVA(257-264) on neighboring bystander Kb-expressing antigen-presenting cells after BALB/c (H-2d) DC phagocytosed E. coli expressing Crl-OVA could not be detected. Together, these data support a cytosolic MHC-I presentation pathway for OVA(257-264) processed from E. coli expressing Crl-OVA by bone marrow-derived DC.