Identification of surface residues of the monocyte chemotactic protein 1 that affect signaling through the receptor CCR2

The CC chemokine, monocyte chemotactic protein, 1 (MCP-1) functions as a major chemoattractant for T-cells and monocytes by interacting with the seven-transmembrane G protein-coupled receptor CCR2. To identify which residues of MCP-1 contribute to signaling though CCR2, we mutated all the surface-exposed residues to alanine and other amino acids and made some selective large changes at the amino terminus. We then characterized the impact of these mutations on three postreceptor pathways involving inhibition of cAMP synthesis, stimulation of cytosolic calcium influx, and chemotaxis. The results highlight several important features of the signaling process and the correlation between binding and signaling: The amino terminus of MCP-1 is essential as truncation of residues 2-8 ([1+9-76]hMCP-1) results in a protein that cannot stimulate chemotaxis. However, the exact peptide sequence may be unimportant as individual alanine mutations or simultaneous replacement of residues 3-6 with alanine had little effect. Y13 is also important and must be a large nonpolar residue for chemotaxis to occur. Interestingly, both Y13 and [1+9-76]hMCP-1 are high-affinity binders and thus affinity of these mutants is not correlated with ability to promote chemotaxis. For the other surface residues there is a strong correlation between binding affinity and agonist potency in all three signaling pathways. Perhaps the most interesting observation is that although Y13A and [1+9-76]hMCP are antagonists of chemotaxis, they are agonists of pathways involving inhibition of cAMP synthesis and, in the case of Y13A, calcium influx. These results demonstrate that these two well-known signaling events are not sufficient to drive chemotaxis. Furthermore, it suggests that specific molecular features of MCP-1 induce different conformations in CCR2 that are coupled to separate postreceptor pathways. Therefore, by judicious design of antagonists, it should be possible to trap CCR2 in conformational states that are unable to stimulate all of the pathways required for chemotaxis.     

Evaluation of anti-pyretic potential of Jussiaea suffruticosa L. extract in rats.

A study was carried out to evaluate the anti-pyretic potential of the methanol extract of the aerial part of Jussiaea suffruticosa Linn. (MEJS) on normal body temperature and yeast-induced pyrexia in albino rats. Yeast suspension (10 ml/kg body wt.) increased rectal temperature after 19 hours of subcutaneous injection. The MEJS, at doses of 100, 200 and 300 mg/kg body wt. p.o., showed significant reduction in normal body temperature and yeast-provoked elevated temperature in a dose-dependent manner. The effect also extended up to 5 hours after the drug administration. The anti-pyretic effect of MEJS was comparable to that of paracetamol (150 mg/kg body wt, p.o.), a standard anti-pyretic agent.     

DNA fingerprinting in the criminal justice system: an overview

DNA fingerprinting is a powerful technology that has revolutionized forensic science. No two individuals can have an identical DNA pattern except identical twins. Such DNA-based technologies have enormous social implications and can help in the fight against crime. This technology has experienced many changes over time with many advancements occurring. DNA testing is a matter of serious concern as it involves ethical issues. This article describes various trends in DNA fingerprinting and the current technology used in DNA profiling, possible uses and misuses of DNA databanks and ethical issues involved in DNA testing. Limitations and problems prevailing in this field are highlighted.     

Role of ecosystem components in Cd removal process of aquatic ecosystem

An experiment was conducted using 15 glass aquariums to ascertain the pathways of removal of cadmium through numerical and compositional manipulation of ecosystem components and their role in Cd removal in different aquatic ecosystems. Each aquarium was provided with surface sediment @ 2 kg, filled with 15 L tap water and randomly distributed into five treatments having three replicates in each. Cadmium chloride (CdCl₂) of analytical grade was added @ 2 mg/L to the water of each aquarium and mixed gently. Except for the first one, the other four systems received unio (Lamellidens marginalis, 55 ± 2.5 g) @ 6 pieces/aquarium. Tilapia (Oreochromis mossambicus, 35 ± 3 g) was introduced @ 6 fish/aquarium in the third and fifth systems, whereas pistia (Pistia stratiotes) was introduced @ 50 g/aquarium in the fourth and fifth systems for a 28-day observation period. The samples of water, sediment, unio, fish and pistia were collected from different systems at 7-day intervals and analyzed. Results revealed that mean substantial reduction of Cd in water varied between 1.820 and 1.994 mg/L in different simulated ecosystems. Ecosystem efficiency of Cd removal varied in the different ecosystems and showed highest (11%) value in the ecosystem carrying five components, which suggested a cumulative effect of increasing number of components employed in different simulated aquatic ecosystems significantly facilitated the reduction of the level of Cd concentration in water column. Pistia exerted (12.88–547.5 times) higher rate of Cd accumulation over the other components employed in five simulated ecosystems of various component structures. Therefore, in the present study, it may be concluded that ecosystems carrying five components exhibited the best performance for optimum minimization of Cd removal from water column. It can also be concluded that ecosystem components showed a variable performance and pistia was the efficient component from the perspective of Cd removal.     

Phenylpropanoid compounds and disease resistance in transgenic tobacco with altered expression of L-phenylalanine ammonia-lyase

Tobacco plants over-expressing L-phenylalanine ammonia-lyase (PAL(+)) produce high levels of chlorogenic acid (CGA) and exhibit markedly reduced susceptibility to infection with the fungal pathogen Cercospora nicotianae, although their resistance to tobacco mosaic virus (TMV) is unchanged. Levels of the signal molecule salicylic acid (SA) were similar in uninfected PAL(+) and control plants and also following TMV infection. In crosses of PAL(+) tobacco with tobacco harboring the bacterial NahG salicylate hydroxylase gene, progeny harboring both transgenes lost resistance to TMV, indicating that SA is critical for resistance to TMV and that increased production of phenylpropanoid compounds such as CGA cannot substitute for the reduction in SA levels. In contrast, PAL(+)/NahG plants showed strongly reduced susceptibility to Cercospora nicotianae compared to the NahG parent line. These results are consistent with a recent report questioning the role of PAL in SA biosynthesis in Arabidopsis, and highlight the importance of phenylpropanoid compounds such as CGA in plant disease resistance.     

Nucleotide binding affinities of the intact proton-translocating transhydrogenase from Escherichia coli

Transhydrogenase (E.C. 1.6.1.1) couples the redox reaction between NAD(H) and NADP(H) to the transport of protons across a membrane. The enzyme is composed of three components. The dI and dIII components, which house the binding site for NAD(H) and NADP(H), respectively, are peripheral to the membrane, and dII spans the membrane. We have estimated dissociation constants (K(d) values) for NADPH (0.87 microM), NADP(+) (16 microM), NADH (50 microM), and NAD(+) (100-500 microM) for intact, detergent-dispersed transhydrogenase from Escherichia coli using micro-calorimetry. This is the first complete set of dissociation constants of the physiological nucleotides for any intact transhydrogenase. The K(d) values for NAD(+) and NADH are similar to those previously reported with isolated dI, but the K(d) values for NADP(+) and NADPH are much larger than those previously reported with isolated dIII. There is negative co-operativity between the binding sites of the intact, detergent-dispersed transhydrogenase when both nucleotides are reduced or both are oxidized.     

Multivessel coronary thrombosis secondary to cocaine use successfully treated with multivessel primary angioplasty

Cocaine use has been associated with a significant risk of myocardial ischemia and myocardial infarction (MI). The previous approach to the treatment of cocaine-induced MI focused on medical treatment with verapamil, nitroglycerine and thrombolytics. Percutaneous revascularization for the cocaine-associated MI has been reported and is the preferred treatment modality. Identification of culprit vessel in the patients presenting with acute myocardial infarction associated with cocaine use is problematic owing to the frequent presence of baseline electrocardiogram (ECG) changes. Chronic cocaine use predisposes to diffuse coronary vasculopathy and may cause systemic alteration of coagulation parameters. Multivessel coronary thrombosis presenting as myocardial infarction associated with cocaine use has not been previously reported. This study describes a case of multivessel coronary thrombosis caused by cocaine ingestion successfully treated with multivessel primary angioplasty.     

Clofarabine in adult acute leukemias: clinical success and pharmacokinetics

Clofarabine is a deoxyadenosine analog synthesized with the intention of retaining the favorable mechanistic properties of fludarabine and cladribine while eliminating their undesirable characteristics. Phase I studies among 32 patients with acute leukemia defined a maximum tolerated dose (MTD) of 40 mg/m2/d given as a one hour infusion daily for 5 days. The dose limiting toxicity (DLT) was transient hepatotoxicity. In a phase II study, 62 patients with acute leukemias received clofarabine at the MTD over 1 hour daily for 5 days. Twenty patients (32%) achieved complete response (CR), 1 had a partial response (PR), and 9 had a CR but without platelet recovery (CRp), for an overall response rate of 48%. Pharmacokinetic studies in the phase I trial revealed marked heterogeneity in peak levels of clofarabine among patients at the end of infusion, however; there was a linear, dose dependent increase in clofarabine concentration in the plasma. Pharmacodynamically, at the MTD, DNA synthesis was inhibited by more than 80% at the end of infusion. In phase II studies, the relationship between the pharmacokinetics of clofarabine triphosphate accumulation and clinical response at the MTD was explored, revealing an accumulation advantage of the cytotoxic triphosphate in leukemia cells of responders. The circulating leukemia blasts of patients who respond to clofarabine therapy exhibited a favorable pharmacokinetic profile. In conclusion, clofarabine is an active agent in the treatment of acute leukemias and MDS, and cellular pharmacokinetics has prognostic significance.