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N Raghavan

Publications and source records attributed to N Raghavan.

At least 19 recordsLinked to original sources
PubMed2006

On methods for gene function scoring as a means of facilitating the interpretation of microarray results.

As gene annotation databases continue to evolve and improve, it has become feasible to incorporate the functional and pathway information about genes, available in these databases into the analysis of gene expression data, for a better understanding of the underlying mechanisms. A few methods have been proposed in the literature to formally convert individual gene results into gene function results. In this paper, we will compare the various methods, propose and examine some new ones, and offer a structured approach to incorporating gene function or pathway information into the analysis of expression data. We study the performance of the various methods and also compare them on real data, using a case study from the toxicogenomics area. Our results show that the approaches based on gene function scores yield a different, and functionally more interpretable, array of genes than methods that rely solely on individual gene scores. They also suggest that functional class scoring methods appear to perform better and more consistently than overrepresentation analysis and distributional score methods.

Databases, Genetic↗
PubMed2001

Differential gene expression in haemocytes of the snail Biomphalaria glabrata: effects of Schistosoma mansoni infection.

Parasite encapsulation and destruction in Biomphalaria glabrata has been shown to involve the cellular component of the snail's internal defence system, the haemocytes. To identify genes involved in the immunobiology of these cells, we used the method of differential display reverse transcriptase polymerase chain reaction (DDRT-PCR) to investigate differential gene regulation in haemocytes isolated from Schistosoma mansoni exposed and unexposed snails. RNA isolated from circulating haemocytes from resistant snails (BS-90 stock), previously exposed to S. mansoni, was analysed using 12 different arbitrary primers in conjunction with an anchored Oligo d(T(11)CG) primer. Transcription profiles between haemocytes of parasite exposed and unexposed snails were compared and a total of 87 differentially regulated bands were identified and isolated. Of these, 65 bands were cloned and used as probes in Southern blots to show the presence of corresponding sequences in the snail genome. RT-PCR was performed to verify the regulation of these transcripts. DNA sequence analysis showed that the majority of the cloned sequences were novel, although a few showed a high degree of sequence similarity to other sequences in the DNA and protein databases. One of these included a differentially expressed transcript that showed a significant degree of sequence identity to E. coli transposase Tn5, an enzyme whose activity is normally associated with generating mobility and instability in the genome.

Amino Acid Sequence↗
PubMed2000

The Wuchereria bancrofti orthologue of Brugia malayi SXP1 and the diagnosis of bancroftian filariasis.

The gene encoding the Wuchereria bancrofti orthologue of the Brugia malayi-derived diagnostic antigen SXP1 was identified from a W. bancrofti L3 cDNA library and characterized. The Wb-sxp-1 cDNA encoded a basic protein with a calculated molecular mass of 20.8 kDa. Wb-SXP-1 was 85% identical to the SXP1 protein described from B. malayi (Bm-SXP-1). The Wb-SXP-1 sequence also showed significant identity with proteins described from B. pahangi, Onchocerca volvulus, Acanthochilonema vitea, Ascaris suum, Loa loa, Litomosoides sigmodontis and Caenorhabditis elegans. The presence of a number of invariant and conserved residues in all of these nematode-derived molecules suggests that Wb-SXP-1 is a member of a new protein family. A recombinant form of Wb-SXP-1 was produced and it was determined that the anti-Wb-SXP-1 antibody response in patients with W. bancrofti infections was restricted to the IgG4 subclass. An anti-Wb-SXP-1 IgG4 ELISA was developed and this assay was found to be 100% sensitive for patients with patent W. bancrofti infection. Sera from individuals experiencing chronic pathology, endemic normals or patients with non-filarial nematode infections had no detectable IgG4 against Wb-SXP-1. While patients with patent Onchocerca volvulus infections were uniformly negative in the Wb-SXP-1 assay, 40% of sera from patent Loa loa infections were positive. When Bm-SXP-1 was used as the antigen under identical conditions, the assay was 88% specific for patent W. bancrofti infections and the antigen was recognized by antibodies from both O. volvulus and L. loa infections. The results strongly suggested that, for certain diagnostic filarial antigens, the use of same-species molecules can enhance the specificity of diagnostic tests.

Amino Acid Sequence↗
PubMed2000

Cost utility analysis of radiographic screening for an orbital foreign body before MR imaging.

BACKGROUND AND PURPOSE: Our purpose was to evaluate the cost-effectiveness of clinical versus radiographic screening for an orbital foreign body before MR imaging. METHODS: Costs of screening were determined on the basis of published reports, disability rating guides, and a practice survey. Base case estimates were derived from published guidelines. A single-state change model was constructed using social cost as the unit of analysis. Sensitivity analysis was performed for each variable. The benefit of screening was avoidance of immediate, permanent, nonameliorable, unilateral blindness. RESULTS: Using base case estimates and a discount rate of zero, we calculated the cost of the current guideline as $328,580 per quality-adjusted life-year saved. Sensitivity analysis identified screening cost as a critical variable. Discount rates and effectiveness of foreign body removal also were found to be important factors. Probability of injury and prevalence of foreign body may impact the analysis. CONCLUSION: Clinical screening before radiography increases the cost-effectiveness of foreign body screening by an order of magnitude, assuming base case ocular foreign body removal rates. Asking the patient "Did a doctor get it all out?" serves this purpose. Occupational history by itself is not sufficient to mandate radiographic orbital screening. Current practice guidelines for foreign body screening should be altered.

Adult↗
PubMed1999

Developmentally regulated expression of a unique small heat shock protein in Brugia malayi.

A screen of an expression library from the fourth larval stage (L4) of the parasitic nematode Brugia malayi resulted in the identification of a 727 bp full-length cDNA with 29-40% identity to members of the small heat shock family of proteins (Bm-hsp-s1). The open reading frame encoded a protein of approximately 18 kDA (Bm-HSP-s1). An alignment of the Bm-HSP-s1 sequence with the sequences of small HSPs from vertebrate and invertebrate species demonstrated that a majority of the identity was concentrated in the central alpha-crystallin domain. Bm-HSP-s1 was constitutively produced by L4 and adult parasites and at low levels by third-stage larvae (L3), but not by first-stage larvae (microfilariae). In adult parasites, Bm-HSP-s1 was localized to the body wall muscle cells and to the cells of the hypodermis/lateral cord. Bm-HSP-s1 production was induced in adult and L3 incubated at 42 degrees C and in L3s during the developmental transition from vector-stage to vertebrate-stage parasites at 37 degrees C. Neither increased nor decreased temperatures induced Bm-HSP-s1 production in microfilariae. Nitric oxide induced low-level, transient Bm-HSP-s1 synthesis in adults, but not in microfilariae. Bm-HSP-s1 did not function as a molecular chaperone to prevent heat-induced aggregation of a test substrate. The developmentally regulated expression and inducable nature of Bm-HSP-s1 suggests that it may have a stage-restricted role in maintaining parasite homeostasis.

Amino Acid Sequence↗
PubMed1998

The transition zone above a lumbosacral fusion.

STUDY DESIGN: The clinical and radiographic effect of a lumbar or lumbosacral fusion was studied in 42 patients who had undergone a posterolateral fusion with an average follow-up of 22.6 years. OBJECTIVE: To examine the long-term effects of posterolateral lumbar or lumbosacral fusion on the cephalad two motion segments (transition zone). SUMMARY OF BACKGROUND DATA: It is commonly held that accelerated degeneration occurs in the motion segments adjacent to a fusion. Most studies are of short-term, anecdotal, uncontrolled reports that pay particular attention only to the first motion segment immediately cephalad to the fusion. METHODS: Forty-two patients who had previously undergone a posterolateral lumbar or lumbosacral fusion underwent radiographic and clinical evaluation. Rate of fusion, range of motion, osteophytes, degenerative spondylolisthesis, retrolisthesis, facet arthrosis, disc ossification, dynamic instability, and disc space height were all studied and statistically compared with an age- and gender-matched control group. The patient's self-reported clinical outcome was also recorded. RESULTS: Degenerative changes occurred at the second level above the fused levels with a frequency equal to those occurring in the first level. There was no statistical difference between the study group and the cohort group in the presence of radiographic changes within the transition zone. In those patients undergoing fusion for degenerative processes, 75% reported a good to excellent outcome, whereas 84% of those undergoing fusion for spondylolysis or spondylolisthesis reported a good to excellent outcome. CONCLUSION: Radiographic changes occur within the transition zone cephalad to a lumbar or lumbosacral fusion. However, these changes are also seen in control subjects who have had no surgery.

Adult↗
PubMed1998

Thioredoxin peroxidases from Brugia malayi.

Parasite-derived antioxidant proteins have been implicated in playing an important role in protection against the oxygen radicals that are generated during aerobic metabolism and in defense against host immune cell attack. Here we report that filarial nematodes include the thioredoxin peroxidase/thiol-specific antioxidant (TPx/TSA) family of antioxidant proteins as part of their complex defense against radical-mediated damage. At the protein level, the TPx/TSA from Brugia malayi (Bm-TPx-1) was approximately 50% identical and approximately 60% similar to TPx/TSAs from mammals, amphibians and yeast. Bm-TPx-1 was also approximately 60% identical to putative TPx proteins from a related filarial nematode, Onchocerca volvulus, and from the free-living nematode Caenorhabditis elegans. That B. malayi may express multiple forms of molecules with TPx/TSA activity was indicated by the identification of a B. malayi gene encoding a second, distinct member of the TPx/TSA family (Bm-tpx-2). Bm-tpx-1 was found to be transcribed in all stages of the parasite present in the mammalian host and the 25 kDa translation product was present in all of the developmental stages studied. The results of immunohistochemical, immunofluorescent and immunoprecipitation studies showed Bm-TPx-1 to be localized in the cells of the hypodermis/lateral chord in adult parasites and not to be present at the surface or in excretory/secretory products. The distribution in the parasite suggests that Bm-TPx-1 may play its major role in countering radicals produced within cells. A recombinant form of Bm-TPx-1 was biologically active and capable of protecting DNA from oxygen radical-mediated damage. Thioredoxin peroxidases may prove to be a critical component in the parasite's defense against injury caused by oxygen radicals derived from endogenous and exogenous sources.

Amino Acid Sequence↗
PubMed1998

Filarial nematode parasites secrete a homologue of the human cytokine macrophage migration inhibitory factor.

Filarial nematode parasites establish long-term chronic infections in the context of an antiparasite immunity that is strongly biased toward a Th2 response. The mechanisms that lead to this Th2 bias toward filarial antigens are not clear, but one possibility is that the parasites produce molecules that have the capacity to proactively modify their immunological environment. Here we report that filarial parasites of humans secrete a homologue of the human proinflammatory cytokine macrophage migration inhibitory factor (MIF) that has the capability of modifying the activity of human monocytes/macrophages. A cDNA clone isolated from a Brugia malayi infective-stage larva expression library encoded a 12.5-kDa protein product (Bm-MIF) with 42% identity to human and murine MIF. MIF homologues were also found to be expressed in the related filarial species Wuchereria bancrofti and Onchocerca volvulus. Bm-mif was transcribed by adult and larval parasites, and the protein product was found in somatic extracts and in the parasite's excretory-secretory products. Immunohistocytochemistry revealed that Bm-MIF was localized to cells of the hypodermis/lateral chord, the uterine wall, and larvae developing in utero. Unexpectedly, the activities of recombinant Bm-MIF and human MIF on human monocytes/macrophages were found to be similar. When placed with monocytes/macrophages in a cell migration assay, Bm-MIF inhibited random migration. When placed away from cells, Bm-MIF induced an increase in monocyte/macrophage migration that was specifically inhibited by neutralizing anti-Bm-MIF antibodies. Bm-MIF is the first demonstration that helminth parasites produce cytokine homologues that have the potential to modify host immune responses to promote parasite survival.

Amino Acid Sequence↗
PubMed1996

Genes expressed in Brugia malayi infective third stage larvae.

We have used a tag sequencing approach to survey genes expressed in the third stage infective larvae of the human filarial nematode parasite Brugia malayi. RNA was isolated from late vector-stage L3 larvae after days 9 or 10 of infection in mosquitos, and converted to cDNA by reverse transcriptase. Double-stranded cDNA was produced by either conventional methods (non-SL cDNA library) or by PCR using the nematode spliced leader (SLI) and oligo(dT) primers (SL cDNA library). Two clone libraries (one from SL and one from non-SL cDNAs) were constructed in lambda ZapII. A set of these full-length clones was selected and 596 inserts were sequenced from the 5' end. We have identified 364 B. malayi genes (the majority of which are new) that encode housekeeping proteins, structural proteins, proteins of immediate immunological or drug-discovery interest as well as a large class of novel sequences which may prove to have significant involvement in host invasion. Extensive, genome-wide approaches to the analysis of larval gene expression are now possible for B. malayi. We present several examples of this approach.

Amino Acid Sequence↗
PubMed1995

Nucleoside diphosphate kinase from the parasitic nematode Brugia malayi.

Using a reverse transcription-polymerase chain reaction (RT-PCR) procedure that exploited the presence of a conserved 22-nucleotide spliced leader (SL) sequence that is trans-spliced to the 5' end of nematode transcripts, a novel Brugia malayi (Bm) infective-stage SL cDNA expression library was constructed and characterized. The library was immunoscreened with rabbit anti-infective-stage antibodies (Ab) and an immunodominant clone, BmG4-7, was identified and characterized. BmG4-7 contained a full-length cDNA that had significant sequence similarity to nucleoside diphosphate kinase (NDK)-encoding sequences reported from a number of species, including Drosophila melanogaster and humans. BmNDK was found to be constitutively transcribed during all stages of parasite development. An anti-BmNDK Ab was used to immunostain a Western blot of extracts from adult and larval parasites. The Ab specifically recognized a 17.5-kDa molecule in all of the parasite extracts. Molecular modeling of the BmNDK showed several regions surrounding the conserved catalytic site that may be important in the design of drugs specific for the disruption of NTP synthesis in filarial parasites.

Amino Acid Sequence↗
PubMed1995

Evaluation of meniscal tears: fast spin-echo versus conventional spin-echo magnetic resonance imaging.

RATIONALE AND OBJECTIVES: We compared the performance of fast spin-echo (FSE) with conventional spin-echo (CSE) magnetic resonance (MR) imaging sequences in the detection of meniscal tears. METHODS: Seventy-three patients underwent MR examination of the knee for suspected internal derangement. Each patient was scanned with a CSE sequence and one of two FSE sequences. The primary difference between the two FSE sequences consisted of the echo train length (ETL). Thirty-seven patients (group 1) were scanned with the FSE I sequence (ETL = 8), and 36 patients (group 2) were scanned with the FSE II sequence (ETL = 4). Menisci were graded as torn or not torn on the basis of their MR appearance. The sequences were compared with each other and with the surgical findings in 31 patients who underwent arthroscopy. RESULTS: In group 1, there was a significant discrepancy between the CSE and FSE I techniques (p = .006). The FSE I sequence detected only 11 of 19 surgically proven torn menisci as opposed to 18 of 19 detected with the CSE sequence. The FSE II sequence performed significantly better in group 2, with an accuracy equal to that of the CSE sequence. CONCLUSION: FSE sequences are extremely technique dependent with regard to detecting meniscal tears and should not replace CSE sequences in this setting until further studies are performed to optimize the technique.

Adolescent↗
PubMed1994

Simultaneous quantification of an anti-inflammatory compound (DuP 697) and a potential metabolite (X6882) in human plasma and urine by high-performance liquid chromatography.

A high-performance liquid chromatographic (HPLC) method using fluorescence detection has been developed for the simultaneous analysis of low nanogram concentrations of an anti-inflammatory drug, 5-Bromo-2-(4-fluorophenyl)-3-[4-(methylsulfonyl)phenyl]thiophene (DuP 697), and a potential metabolite (X6882) in human plasma and of DuP 697 in human urine. This assay method used an EM Separations Lichrospher C18 endcapped column. The mobile phase was acetonitrile-water (75:25, v/v). The detection of DuP 697 and X6882 was by fluorescence at excitation and emission wavelengths of 256 and 419 nm, respectively. The chromatographic system could separate DuP 697 from X6882, the external standard (anthracene), and other endogenous substances present in human plasma. In human plasma the limits of quantification for DuP 697 and X6882 were 3 and 20 ng/ml, respectively; the limit of quantification for DuP 697 in human urine was 5 ng/ml. These compounds were shown to be stable in frozen (-20 degrees C) human plasma and urine for at least 9 weeks. The assay described has been used to characterize DuP 697 pharmacokinetics after oral administration in humans.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1994

Cloning and characterization of a potentially protective chitinase-like recombinant antigen from Wuchereria bancrofti.

While there is no direct evidence demonstrating the existence of protective immunity to Wuchereria bancrofti infection in humans, the presence of individuals, in populations in areas where infection is endemic, with no clinical evidence of past or current infection despite appreciable exposure to the infective larvae, suggests that protective immunity to filarial parasites may occur naturally. Earlier work indicated that such putatively immune individuals generated antibodies to a 43-kDa antigen from larval extracts of the related filarial parasite Brugia malayi that was recognized by only 8% of the infected population. With rabbit antiserum raised against this 43-kDa antigen, this current study identified a recombinant clone, WbN43, with an insert size of 2.3 kb, from a W. bancrofti genomic expression library. The recombinant fusion protein was differentially recognized by the putatively immune individuals but not by the infected patients. The coding sequence (684 bp) from the 5' end had significant sequence similarity to chitinases from Serratia marcescens, Bacillus circulans, Streptomyces plicatus, and B. malayi. Peptide sequencing of the expressed product also defined a chitinase-like sequence. Molecular characterization indicated WbN43 to be a low-copy-number gene, with expression predominantly in infective larvae and microfilariae but not in adult parasites.

Adult↗
PubMed1993

Seroprevalence of infectious bursal disease (IBD) in parts of Tamil Nadu, India.

Sera samples of 400 apparently healthy broiler poultry from 23 commercial farms and 120 sera samples from commercial layer poultry [with specific lesions of infectious bursal disease (IBD) in the internal organs] were screened for the presence of IBD serotypes I, II and the variant strain by the agar gel immuno diffusion (AGID) test, employing standard antigens. Likewise the bursae, spleen and kidney specimen from 96 layer poultry with lesions suggestive of IBD were screened for the presence of IBD antigens employing standard antisera by the AGID test. The study revealed that antibodies to all three strains of IBD virus existed in 73.75% of sera from 400 broilers, while only serotype II antigen was detectable in tissues of the 96 layer birds that exhibited characteristic lesions of IBD. It was concluded that the serotype I vaccine did not protect the layer poultry against serotype II infection and the totally unvaccinated broiler poultry had suffered subclinical infection from all the three serotypes of IBD virus.

Animals↗
PubMed1992

Identification of a repetitive element in the snail Biomphalaria glabrata: relationship to the reverse transcriptase-encoding sequence in LINE-1 transposons.

BamHI-digested Biomphalaria glabrata DNA contains a repetitive 2.0-kb fragment which is readily discernible by ethidium bromide staining. We present evidence that this repetitive element is related at both the nucleotide and amino acid levels to long interspersed nuclear element (LINE)-like transposons. Although comparable elements have been described in several invertebrates, this is the first report of a molluscan homologue. In common with LINE transposons, an open reading frame in the B. glabrata element shows significant homology to reverse transcriptase--a feature believed to allow the dissemination of these elements in the eukaryotic genome.

Amino Acid Sequence↗
PubMed1992

Characterization of a muscle-associated antigen from Wuchereria bancrofti.

A recombinant clone, WbN1, isolated from a genomic expression library of Wuchereria bancrofti and showing restricted specificity at the DNA level (Southern and PCR analyses) for Wuchereria bancrofti and Brugia malayi has been previously described. Sequence analysis of WbN1 indicated that it had notable similarity to myosin. Further characterization using in situ hybridization has localized the mRNA in the muscle of the adult parasite and in the microfilariae. Rabbit polyclonal antiserum, raised against the recombinant WbN1 fused to the maltose-binding protein, recognized a 200-kDa polypeptide in immunoblots containing B. malayi antigen extracts. The same antibody also recognized myosin extracted from Brugia pahangi, Onchocerca volvulus, and Caenorhabditis elegans. Localization using the rabbit antiserum revealed the presence of the antigen in the adult muscle tissue and in the microfilariae; the same antibody inhibited the binding of a monoclonal antibody 28.2 (directed toward MHC B of C. elegans myosin) to the recombinant WbN1 antigen and also to purified C. elegans myosin. Based on homology data, structural location, competitive ELISA, and immunoblot we conclude that WbN1 is related to myosin or a similar myofibrillar protein.

Amino Acid Sequence↗