The impact of multiple stenosis and aneurysms on arterial diseases: A cardiovascular study.

The comparative effect of serial stenosis and aneurysms arteries on blood flow is examined to identify atherosclerotic diseases. The finite element approach has been used to solve the continuity, momentum, and Oldroyd-B partial differential equations to analyze the blood flow. Newtonian and non-Newtonian both cases are taken for the viscoelastic response of blood. In this study, the impact of multiple stenotic and aneurysmal arteries on blood flow have been studied to determine the severity of atherosclerosis diseases through the analysis of blood behavior. The novel aspect of the study is its assessment of the severity of atherosclerotic disorders for the occurrence of serial stenosis and aneurysm simultaneously in the blood vessel wall in each of the four cases. The maximum abnormal arterial blood flow effect is found for the presence of serial stenoses compared to aneurysms which refers to the severity of atherosclerosis. At the hub of stenosis, the blood velocity magnitude and wall shear stress (WSS) are higher, whereas the arterial wall normal gradient values are lower. For all cases, the contrary results are observed at the hub of the aneurysmal model. The blood flow has been affected significantly by the increases in Reynolds number for both models. The influence of stenotic and aneurysmal arteries on blood flow is graphically illustrated in terms of the velocity profile, pressure distribution, and WSS. Medical experts may use this study's findings to assess the severity of cardiovascular diseases.

Health Related Quality of Life in Patients with Pulmonary Tuberculosis.

Tuberculosis (TB) is one of the major public health issues in many developing nations especially in Bangladesh. Though most focus is being directed towards mortality and incidence rate, the changes in morbidity and other health status parameters are not been well considered. The aim of the study was a comprehensive assessment of patients with pulmonary tuberculosis by measuring patient's quality of life which may lead to better outcome in patients' health, infection surveillance and prevention programs. This prospective study was conducted in the department of Respiratory and Internal Medicine, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh from September 2015 to March 2017. The quality of life scores of 61 smear positive pulmonary tuberculosis cases were measured by validated Bangla version of SF-36 questionnaire before or at the starting of treatment, after the initial phase and at the end of treatment. Then the score was compared with those of 75 healthy matched controls. The changes of the quality of life with the stage of treatment and with socio-demographic variables were assessed. Before treatment, all domains of HRQoL of the pulmonary TB patients were significantly lower than those of the control group (p<0.001). At the end of six-month treatment period, HRQoL of the pulmonary TB patient had significantly increased compared to before treatment (p<0.001). There was no significant difference of scores after six months of treatment with that of control (p>0.05). The lowest score in tuberculosis patients was related to general health perception and vitality. Patients with low socio economic status, low educational level, prolonged disease duration and increased number of symptoms had lower HRQoL scores.

Etiological Evaluation of Patients with Lymphadenopathy by Clinical, Histopathological and Microbiological Assessment.

Lymph node enlargement is a common presenting complaint in outpatient and inpatient department. The present observational cross sectional study was conducted in department of Internal Medicine, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh during the period from December 2014 to May 2016 to evaluate etiologies of significant lymphadenopathy by clinical, histopathological and microbiological assessment. Biopsy/FNA materials of 177 patients of 18-75 years age range with significant lymphadenopathy were sent for histopathology/cytology, Gram stain & culture, AFB stain & culture and Gene Xpert. Among them, 102(57.62%) were granulomatous lymphadenitis, 52(29.38%) were lymphoma, 12(6.78%) reactive lymphadenitis, 7(3.95%) metastatic malignancy, 2(1.13%) atypical lymphoid hyperplasia, 1(0.57%) myeloid sarcoma and 1(0.57%) chronic sialadenitis. Growth of MTB was on 23(22.55%) cases; among 102 granulomatous lymphadenitis and Gene Xpert was positive in 73(71.56%) cases with 100% Rif. sensitive. Gene Xpert is an important tool for diagnosis of tuberculous lymphadenitis. Time of symptoms to diagnosis of most of the TBL patients was within 2-8 months.

Serum Zinc Concentrations in Patients with Pulmonary Tuberculosis.

Bangladesh is a tuberculosis (TB) burden country. It is one of the most important causes of mortality and morbidity and a major barrier of social and economic development. Zinc is a major trace element and an essential component of the body immune system. It's an important determinant of resistance to infection by maintaining cell mediated immunity. This analytical case control study was conducted in the Department of Internal Medicine, Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh to see the association of serum zinc concentrations with pulmonary tuberculosis (PTB) in adult population (18-60 years) from January 2015 to January 2016. Freshly diagnosed PTB patients before initiating anti-TB chemotherapy as cases (N=43) and TB negative subjects as controls (N=48) were included conveniently in this study with a rigid selection criteria. Serum zinc concentrations were estimated by using atomic absorption spectrophotometer. The mean±SD age and BMI of the case group and control group were 33.30±14.71 and 32.69±11.60 years, 19.88±2.31 and 22.08±2.80 kg/m2 respectively. The concentrations of serum zinc were significantly lower (P=0.01) in PTB group (840.9±230.0 µgm/l) compared with the control group (965.6±219.9 µgm/l). There was marked variation of mean±SD serum zinc concentrations between male (1008.95±246.16 µgm/l) and female (937.24±200.35 µgm/l) in control group (P=0.182) though the variation is minimal in PTB group (P=0.724). The serum zinc concentrations showed positive correlation with BMI (P=0.642) but negative correlation with age (P=0.023) in both case and control. The lower serum zinc concentrations (12.06%) in PTB patients indicate relative immune deficiency. Routine assessment of serum zinc concentration of PTB patients should be considered and further outcome should be assessed with zinc supplementation.

Inducing pluripotency in vitro: recent advances and highlights in induced pluripotent stem cells generation and pluripotency reprogramming.

Induced pluripotent stem cells (iPSCs) are considered patient-specific counterparts of embryonic stem cells as they originate from somatic cells after forced expression of pluripotency reprogramming factors Oct4, Sox2, Klf4 and c-Myc. iPSCs offer unprecedented opportunity for personalized cell therapies in regenerative medicine. In recent years, iPSC technology has undergone substantial improvement to overcome slow and inefficient reprogramming protocols, and to ensure clinical-grade iPSCs and their functional derivatives. Recent developments in iPSC technology include better reprogramming methods employing novel delivery systems such as non-integrating viral and non-viral vectors, and characterization of alternative reprogramming factors. Concurrently, small chemical molecules (inhibitors of specific signalling or epigenetic regulators) have become crucial to iPSC reprogramming; they have the ability to replace putative reprogramming factors and boost reprogramming processes. Moreover, common dietary supplements, such as vitamin C and antioxidants, when introduced into reprogramming media, have been found to improve genomic and epigenomic profiles of iPSCs. In this article, we review the most recent advances in the iPSC field and potent application of iPSCs, in terms of cell therapy and tissue engineering.

Chronic calcific pancreatitis and pancreatic cancer.

An observational cross-sectional study of 50 cases of chronic calcific pancreatitis patients was conducted in Bangabandhu Sheikh Mujib Medical University (BSMMU) and some other tertiary level hospitals of Dhaka city from August 2008 to July 2010. Patients required laparotomy for different modalities of surgical treatment to manage chronic calcific pancreatitis were included in the study. Biopsy was taken from panceatic duct containing stone during laparotomy to determine the histopathological changes. Among 50 cases female predominance was observed. Male, female ratio was 2:3. Majority (62%) patients were in 20 to 40 years age group. Female presented earlier than male (20-30 years and 30-40 years respectively). All patients complained recurrent attack of epigastric pain. Other presentations were diabetes (74%), malnutrition and weight loss (56%), steatorrhoea (24%) and jaundice (12%). Adenocarcinoma was found in 3(6%) patients (2 male and 1 female) and rests were chronic pancreatitis. Several studies showed the association between chronic calcific pancreatitis and pancreatic cancer. Further large scale study is required to find out the national incidence level.

Nanostructured transducer surfaces for electrochemical biosensor construction--interfacing the sensing component with the electrode.

For fabrication of effective electrochemical biosensors, interfacing the biomolecular receptor with the underlying transducer represents a critical step. The actual approach taken depends on the tethering layer covering the transducer, which is typically either a conducting polymeric matrix, or a thin film, such as an alkanethiol monolayer. Non-specific immobilisation methods can be either covalent, or non-covalent affinity attachment, with multipoint electrostatic attachment of the sensing biomolecule to either a polyanionic or polycationic layer representing the most common approach. Many specific affinity immobilisation strategies exist, but the majority make use of one of two binding systems. The first relies on the specific and strong affinity between biotin and proteins of the avidin family, with both bioreceptor and transducer bearing pendant biotins and avidin used as the crosslinker. The second approach employs a metal chelating group on the transducer to which can be bound a polyhistidine tag present on the N- or C-terminus of the receptor protein and which can be introduced genetically, when the expression sequence for a recombinant proteins is designed.

The role of neutrophils in LPS-induced changes in pulmonary function in conscious rats.

We have previously reported on a model of lipopolysaccharide (LPS)-induced pulmonary inflammation in rats, where LPS-challenged animals develop a significant pulmonary neutrophilia and mucus hypersecretion. In the current studies, we utilized whole body plethysmography and computer assisted data acquisition to examine changes in pulmonary parameters, e.g. frequency (f) tidal volume and Penh as a measure of bronchoconstriction, due to LPS-challenge in conscious rats. Compared to saline challenge, LPS-challenged rats displayed a significant increase in (f) which began within 30 min, peaked by 2 h and remained elevated up to 24 h. Mirroring this increase in (f) was a decrease in the observed tidal volume of LPS-challenged rats. Additionally, compared to saline challenge, LPS-challenge provoked a significant and spontaneous bronchoconstriction, as measured by Penh, 2 h after challenge. In order to further understand these observed LPS-induced pulmonary changes, we utilized two classes of pulmonary obstructive disease standards, namely, bronchodilators and anti-inflammatory agents, and examined their ability to affect the spontaneous bronchoconstriction and the increase in (f) seen at two discrete time points, i.e. 2 and 24 h after LPS-challenge. While ineffective on either the 2 h increase in (f) or the LPS-induced inflammation, animals pretreated with salbutamol (10 mg/kg, p.o.) were protected from the increase in (f) seen at the 24 h time point after LPS-challenge. In contrast, when animals were pretreated with theophylline (10 mg/kg, p.o.) no effect on the LPS-induced pulmonary inflammation or increase in (f) was noted. Meanwhile, in animals pretreated with either betamethasone (3 mg/kg, p.o.) or SB207499 (10 mg/kg, p.o.), a PDE4 inhibitor, doses previously shown to block the LPS-induced neutrophilic inflammation, the persistent increase in (f) seen at 24 h was attenuated, but neither compound was able to attenuate either the increase in (f) or the spontaneous bronchoconstriction seen at 2 h. In summary, the intra-tracheal LPS-challenge of rats results in pulmonary inflammation and dysfunction, which is similar to that seen in COPD patients. We conclude that the early increase in (f) and bronchoconstriction are not dependent upon airway inflammation, but airway inflammation most likely contributes to the persistent increase in (f) seen at 24 h.

Human phosphodiesterase 8A splice variants: cloning, gene organization, and tissue distribution.

We have cloned cDNAs representing five full-length human phosphodiesterase (PDE) 8A splice variants (PDE8As 1-5) from testis and T cells. PDE8A1 encodes a hydrophilic protein of 829 amino acids, containing an N-terminal REC domain, a PAS domain, and a C-terminal catalytic domain. PDE8A2 encodes a protein of 783 amino acids, identical to PDE8A1 but lacking the PAS domain. PDE8A3 encodes a shorter protein equivalent to the C-terminal 449 amino acids of PDE8A1, containing the catalytic but not the REC and PAS domains. PDE8A4 and PDE8A5, though different from each other at the nucleotide level, encode an identical protein equivalent to the C-terminal 582 amino acids of PDE8A1, including half of the PAS domain. The PDE8A gene is revealed to contain 23 exons, and its exon-intron boundaries have been defined. In addition, we have mapped a common transcription initiation site, and further determined the upstream 5'-flanking sequence of 1740 bp containing the putative promoter. Compared to PDE8A1, PDE8As 2-5 appear to be expressed in much lower abundance. Among various tissues and organs, PDE8A1 and PDE8A2 are expressed at various levels.

Characterization of human, dog and rabbit corpus cavernosum type 5 phosphodiesterases.

Human, dog and rabbit corpus cavernosum type 5 phosphodiesterases (PDE5) were isolated and their characteristics were compared. The three enzymes showed Km values of 0.8, 2.1 and 2.3 uM, respectively. They exhibited similar pH-dependence with optimal pH being 7.5. They required Mg++ for activity and the activity was suppressed by high concentrations of Zn++ (0.1-1 mM). Sildenafil potently inhibited the three enzymes with IC50 values of 3.6, 1.7 and 3.0 nM, respectively. Dipyridamole and IBMX (3-isobutyl-1-methylxanthine) each also inhibited the three enzymes with similar, albeit lower, potencies (IC50 about 1.1 and 5.7 uM, respectively). However, zaprinast exhibited a significantly higher potency against the rabbit enzyme (IC50 53 nM) than against the human and dog PDE5s (IC5s 332 and 217 nM, respectively). Thus, the corpus cavernosum PDE5s are very similar among the various species with the only significant difference being their sensitivity to zaprinast. Human platelet PDE5 was also characterized by comparison with the corpus cavernosum enzyme. The platelet enzyme exhibited a Km, pH-, Mg++- and Zn++-dependence, and sensitivity to sildenafil and zaprinast very similar to those of the corpus cavernosum PDE5. However, compared with corpus cavernosum PDE5, the platelet enzyme exhibited higher sensitivity to dipyridamole and IBMX (IC50 0.46 and 1.8 uM, respectively). This study shows that despite similar kinetics and enzymatic properties, corpus cavernosum PDE5s from different species, and corpus cavernosum and platelet PDE5s, can have differential sensitivity to pharmacological inhibitors.

Cloning, characterization, and tissue distribution of mouse phosphodiesterase 7A1.

We have cloned a cDNA representing mouse phosphodiesterases (PDE) 7A1. The open reading frame encodes a protein of 482 amino acids with a predicted molecular mass of 55417. Like human PDE7A variants, mouse PDE7A1 and A2 are 5' splice variants from a common gene. The distinct N-terminal sequence of mouse PDE7A1 is highly homologous to the corresponding sequence of human PDE7A1 with a similarity of 98% but not to that of mouse PDE7A2 (with a similarity of 12%), and is more hydrophilic than that of mouse PDE7A2. Mouse PDE7A1 expressed in SF9 cells has been compared with human PDE7A1 under identical conditions. Mouse PDE7A1 has a Km for cAMP of 0.2 microM, an optimal pH of 7.5, an IC(50) value of 14 microM for 3-isobutyl-1-methylxanthine (IBMX), and is dependent on Mg(2+) for activity. All these characteristics are very similar to those of human PDE7A1. In mice, PDE7A1 is expressed in tissues of the immune system (lymph node, thymus, spleen, and blood leukocyte), testis, brain, kidney and lung but not in skeletal muscle, heart, embryo, or liver, while PDE7A2 is expressed in skeletal muscle, heart, embryo, and kidney, but not in the other tissues. This tissue distribution profile is very similar to that in humans, and hence suggests that PDE7A1 and 7A2 might play a similar role in different species.

Receptor reserve analysis of the human CCR3 receptor in eosinophils and CCR3-transfected cells.

A novel pharmacological study of CCR3 receptor reserve in a CCR3-transfected cell (CREM3) and human eosinophils was done; functional responses measured were increases in intracellular calcium and chemotaxis. Eotaxin, eotaxin-2, monocyte chemoattractant protein-4 (MCP-4), RANTES, and MCP-3 induced similar maximal eosinophil chemotaxis, whereas MCP-3 and RANTES induced submaximal calcium responses in eosinophils compared to eotaxin, MCP-4, and eotaxin-2. This suggested a receptor reserve in the chemotaxis response. Receptor reserve was quantitated for eotaxin. Occupancy of all CCR3 receptors was required for a maximal calcium response in both CREM3 and eosinophils (reserve = 1.0 or 0.17, respectively); the stimulus-calcium response relationship was linear, indicating no receptor reserve. In contrast, in eosinophils a large receptor reserve (6.5) was found for chemotaxis, where occupancy of 15% receptors drove half-maximal responses. These studies indicate that CCR3 interacts with G-proteins that are poorly coupled to the calcium response, whereas coupling efficiency and/or amplification to the chemotaxis apparatus in human eosinophils is significantly greater.

Mapping and characterization of the epitope(s) of Sch 55700, a humanized mAb, that inhibits human IL-5.

mAb against human IL-5 inhibit pulmonary eosinophilia, tissue damage and airway hyper-reactivity in allergic animal models. Sch 55700 is a humanized, neutralizing anti-IL-5 antibody. To better understand the molecular mechanism by which Sch 55700 blocks IL-5 bioactivity, we have mapped its epitope by scanning IL-5 with synthetic peptides. Those peptides containing a region, ERRRV, corresponding to amino acids 89-93 of IL-5 specifically interact with both Sch 55700 and its parental rat IgG, 39D10. Among the five residues of this region, all three arginine residues were particularly critical for interaction of these peptides with Sch 55700. We further characterized this region by alanine scanning using site-directed mutagenesis. Examination of COS-expressed IL-5 mutants by Western blot showed that single mutations of E(89), R(90), R(91) or R(92) to alanine caused a loss of IL-5 binding to both Sch 55700 and 39D10. We further demonstrated in surface plasmon resonance studies using a BIAcore biosenosor that E(89), R(90) or R(91) are involved in the interaction between IL-5 and its receptor alpha subunit. Based upon the findings here and previously reported structures of the IL-5 and 39D10 variable region, we propose a model suggesting that the molecular interactions between the IL-5 and Sch 55700 mainly involve several ion pair interactions. We conclude that Sch 55700 occupies a region, ERRR, on IL-5 that is essential for its interaction with the receptor and thereby blocks IL-5 bioactivity.

The assignment of chemokine-chemokine receptor pairs: TARC and MIP-1 beta are not ligands for human CC-chemokine receptor 8.

Identification of chemokine receptors and their associated ligands is crucial to the understanding of most immune reactions. Three human chemokines [I-309, thymus and activation-regulated chemokine (TARC) and macrophage inflammatory protein-1beta (MIP-1beta)] have been reported to be ligands for CC-chemokine receptor 8 (CCR8). In this report, we present evidence that TARC and MIP-1beta did not bind to or induce chemotaxis through CCR8 on a stable transfected cell line (1D-21) and did not bind to CCR8 on in vitro differentiated human CD4(+) Th(2) cell cultures. Also, I-309-dependent calcium mobilization in 1D-21 cells and in Th(2) cells was desensitized by I-309 but not by MIP-1beta or TARC. These results provide strong evidence that, at physiologically relevant concentrations, I-309 is the only known human ligand for CCR8. These data also provide a framework for suggesting minimum requirements for the assignment of chemokine receptor-ligand pairs.

Conformational considerations in the design of dual antagonists of platelet-activating factor (PAF) and histamine.

Following the discovery of the first dual antagonist of platelet-activating factor (PAF) and histamine, 1-acetyl-4-(8-chloro-5,6-dihydro-11H-benzo[5,6]cyclohepta[1,2-b]pyridin- 11-ylidene)piperidine, Sch 37370, 1, a related series of structures, exemplified by (+/-)-1-acetyl-4-(8-chloro-5,6-dihydro-11H-benzo[5,6]-cyclohepta[1,2-b] pyridin-11-yl)piperazine, Sch 40338, 2, were prepared. Interestingly, the compounds exhibited a parallel structure antiallergy activity relationship, suggesting that the two series may adopt a common conformation at the PAF receptor. Conformational analysis led to a proposal for this bioactive conformation accessible to both series. The synthesis of novel conformationally constrained analogues that might mimic the proposed bioactive conformation of these compounds, and the evaluation of their in vitro antiallergy activity form the subject matter of this report.

The profiles of human and primate [3H]Nalpha-methylhistamine binding differ from that of rodents.

Characterization of the histamine H3 receptor in rodent species has been extensive but limited characterization has been done with primate or human tissue. We have characterized the binding of [3H]Nalpha-methylhistamine to cynomolgus monkey and human brain membranes to determine whether there are any significant differences among species' pharmacology. In monkey, [3H]Nalpha-methylhistamine bound, in a guanine nucleotide-sensitive fashion, to an apparently homogeneous class of sites at equilibrium (K(D) = 1.4 nM, Bmax = 34 fmol/mg protein). The profile of binding was broadly similar to that of rodents, with a couple of significant differences. Most notably, the potency of the histamine H3-receptor-specific antagonist thioperamide (Ki = 240 nM) was substantially less than reported for rodents and under assay conditions that yield a two-site curve fit in rodents only a single class of thioperamide binding sites was detected in monkey. Burimamide, however, yielded a two-site curve fit (KiH = 6.7 nM, KiL = 1100 nM) independent of the presence of sodium in the assay, as it does in rodents. Characterization of the human brain histamine H3 receptor showed that it was similar to the monkey and not rodent receptor. Our findings indicate that differences between primate and rodent histamine H3 receptors of potentially serious importance for the discovery of antagonists active in humans do exist.

Phosphodiesterase 4B2 is the predominant phosphodiesterase species and undergoes differential regulation of gene expression in human monocytes and neutrophils.

The type 4 phosphodiesterase (PDE4) is the predominant PDE isozyme in various leukocytes and plays a key role in the regulation of inflammatory cell activation. There are four PDE4 subtypes (A, B, C, and D), and within each subtype, there are multiple variants. Very recently, we found in monocytes that PDE4B gene expression is selectively induced by lipopolysaccharide (LPS) and that the induction is inhibited by interleukin (IL)-10 and IL-4. In this study, we show that the PDE4B gene is constitutively expressed in neutrophils and that this expression remains unaffected by LPS or IL-10. PDE4B is the predominant subtype in neutrophils and in unstimulated or LPS-stimulated monocytes, and in these cells, the PDE4B2 variant is the only detectable molecular species of PDE4B. Therefore, PDE4B2 is the predominant PDE isoform in human neutrophils and monocytes, and its expression is regulated differently by these two cell types. Furthermore, leukocytes are the most dominant source of PDE4B2, suggesting that PDE4B2 is a relatively specific target for discovering anti-inflammatory drugs.

A scintillation proximity assay for human interleukin-5 (hIL-5) high-affinity binding in insect cells coexpressing hIL-5 receptor alpha and beta subunits.

The high-affinity receptor for human interleukin-5 (hIL-5) is composed of alpha and beta subunits. A baculovirus expression system was established in Sf9 cells capable of expressing hIL-5 receptor alpha and beta subunits simultaneously. By using wheat germ agglutinin (WGA)-coated scintillation proximity assay (SPA) beads to capture 125I-labeled hIL-5-bound Sf9 cells, a SPA was developed and used to measure hIL-5 high-affinity binding. The hIL-5 receptors expressed in the Sf9 cells represented a single class of high-affinity binding sites with a dissociation constant (Kd) of 0. 24 nM and a density of 2.95 x 10(5) sites/cell. This is the first study in which the high-affinity Kd value similar to that for hIL-5 binding to human eosinophils was achieved using a recombinant expression system. The SPA compared favorably with the filter binding assay with regard to various binding parameters. We also found that several lectins, when coated on SPA beads, were even more effective than WGA-coated SPA beads for capturing the insect cells. We conclude that the baculovirus expression system was highly efficient in producing the high-affinity hIL-5 receptors and that the SPA was a simple and sensitive assay that could be readily adapted into a high-throughput screening format. The SPA described here could be a prototype for binding assays for other multimeric receptors.

Effects of chronic anti-interleukin-5 monoclonal antibody treatment in a murine model of pulmonary inflammation.

The maturation of eosinophils in bone marrow, their migration to pulmonary tissue, and their subsequent degranulation and release of toxic granule proteins contributes to the pathophysiology observed in asthma. Interleukin-5 (IL-5) is essential for these processes to occur. Therefore, much emphasis has been placed on attempts to inhibit the production or activity of IL-5 in order to attenuate the inflammatory aspect of asthma. In this report, the immunological consequences of long-term exposure to an antibody recognizing IL-5 (TRFK-5) were studied in a murine pulmonary inflammation model. A single dose of TRFK-5 (1 mg/ kg, intraperitoneally) reversibly inhibited antigen-dependent lung eosinophilia in mice for at least 12 wk and inhibited the release of eosinophils from bone marrow for at least 8 wk. Normal responses to aerosol challenge were attained after 24 wk. In mice treated acutely with antibody (2 h before challenge), 50% inhibition of pulmonary eosinophilia occurred when 0. 06 mg/kg TRFK-5 was administered (intraperitoneally; ED50), resulting in 230 ng/ml (IC50) in serum. In mice treated with one dose of TRFK-5 (1 mg/kg) and rested before challenge, the antibody exhibited a half-life of 2.4 wk. After 18 to 19 wk, antigen challenge-induced eosinophilia was inhibited by 50% and serum levels of TRFK-5 were 25 ng/ml. TRFK-5 remaining in mice 8 wk after a single injection of TRFK-5 was sufficient to inhibit at least 50% of the eosinophilia induced in blood 3 h after injection of recombinant murine IL-5 (10 microg/kg, intravenously). To assess the biologic effect of long-term exposure of mice to antibody, several parameters of immune-cell function were measured. Throughout the extended period of activity of TRFK-5 (>/= 12 wk) there were no gross effects on antigen-dependent increases in T-cell recruitment into bronchoalveolar fluid (BALF), in IL-4 and IL-5 steady-state mRNA levels in lung tissue, or in immunoglobulin E (IgE) and IgG levels in serum. There was a small increase in IL-5 steady-state mRNA production in TRFK-5-treated mice after 2 h or 2 wk, but this was not observed at other times examined. In untreated mice, IL-5 steady-state mRNA production in response to antigen challenge decreased > 6-fold with age, although at all time points there was an increase in mRNA levels following challenge. Therefore, at later times, 25 ng/ml rather than 230 ng/ml of TRFK-5 inhibited BALF eosinophilia, probably because of reduced IL-5 levels. Twenty-four weeks after treatment with TRFK-5, when challenge-induced eosinophilia was restored, there was an excess of CD4(+) T cells in BALF from challenged mice. However, these T cells had no measurable effects on other responses to challenge, including cytokine production, B-cell accumulation, and immunoglobulin production in serum. Thus, the biologic duration of TRFK-5 was several months, and its activity was due to the presence of antibody above a therapeutic threshold rather than to any profound effect on the immune system.

Phosphodiesterase 4B gene transcription is activated by lipopolysaccharide and inhibited by interleukin-10 in human monocytes.

There are four different genes encoding the cAMP-specific phosphodiesterase (PDE4) isozymes (A, B, C, and D). cAMP has been the only agent known to induce PDE4 gene expression. In the present study, we demonstrate, for the first time, that lipopolysaccharide (LPS) significantly and selectively stimulated PDE4B mRNA production in human monocytes. The LPS stimulation occurred very rapidly (in 30-45 min) and in a dose-dependent manner (0.01-100 ng/ml). We also demonstrate that LPS induction of PDE4B mRNA expression was inhibited strongly by interleukin (IL)-10. The inhibition with IL-10 was dose-dependent (0.1-10 ng/ml). IL-4 also inhibited the LPS induction, but to a lesser extent than IL-10. PDE4B mRNA expression was also stimulated by dibutyryl-cAMP. Interestingly, unlike LPS induction, the dibutyryl-cAMP induction of PDE4B mRNA expression was not inhibited by IL-10. By performing nuclear run-on and mRNA stability assays, we demonstrate further that IL-10 inhibited LPS-stimulated PDE4B mRNA synthesis by abolishing the gene transcription rather than by enhancing mRNA degradation. The present study suggests that PDE4B, as the only LPS-inducible PDE4 subtype, may be an appropriate target for discovering anti-inflammatory drugs.