Fine needle aspiration diagnosis in HIV-related lymphadenopathy in Mangalore, India

OBJECTIVE: To evaluate the usefulness of fine needle aspiration (FNA) study of lymph nodes in HIV-positive patients. STUDY DESIGN: The study was conducted at Kasturba Medical College, Mangalore, India. Samples from lymph nodes of 48 HIV-positive patients were taken and air dried, and wet smears were made. After staining with routine cytologic stains and special stains, detailed cytomorphologic study was conducted. RESULTS: Tuberculosis accounted for nearly half (48%) the cases,followed by HIV lymphadenitis (36%), lymphoma (10%), suppurative lymphadenitis (2%), Mycobacterium avium-intracellulare lymphadenitis (2%) and metastases (2%), in descending order of their frequency. A suppurative picture, which was found in 13% of cases of tuberculous lymphadenitis in AIDS patients, occasionally was misleading without the help of acid fast bacilli stain. CONCLUSION: FNA is a useful tool in the diagnosis of lymphadenopathy in HIV-positive patients provided that proper safety measures are taken to avoid contracting the infection.     

Nitration of profilin effects its interaction with poly (L-proline) and actin

Profilin from bovine spleen was nitrated with peroxynitrite; immunoblotting and spectrophotometric quantitation of nitrotyrosine residues suggested nitration of a single tyrosine residue in profilin with a stoichiometry of 0.6 mol of nitrotyrosine/mole of profilin. A decrease in the nitrotyrosine immunoreactivity of nitroprofilin during digestion with carboxypeptidase Y indicated that nitrotyrosine is located at the C-terminus of profilin. Nitroprofilin interaction with ligands such as phosphatidylinositol 4,5-bisphosphate, actin and poly (l-proline) was analyzed by monitoring the tryptophan fluorescence. Scatchard plot and binding isotherm data obtained revealed no significant difference in affinity of nitroprofilin to phosphatidylinositol 4,5-bisphosphate (K(d) of 4.8 +/- 0.5 muM for profilin, and K(d) of 5.7 +/- 0.6 muM for nitroprofilin), while poly (l-proline) binding studies revealed a twenty-fold increase in the affinity of profilin to poly (l-proline) upon nitration (K(d) of 21.8 +/- 1.7 muM for profilin, and K(d) of 1.1 +/- 0.1 muM for nitroprofilin). Actin polymerization studies involving pyrene-labeled actin indicated that profilin nitration inhibits the actin sequestering property of profilin. The critical actin monomer concentration (C(c)) was 150 and 250 nM in the presence of nitroprofilin and profilin, respectively. Thus, nitric oxide and free radicals produced under different conditions could alter the functions of profilin through nitration, such as its interaction with actin and poly (l-proline).     

Crystal structure of N-(benzyloxycarbonyl)aminoethyl-2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranoside: stabilization of the crystal lattice by a tandem network of N-H. . .O, C-H. . .O, and C-H. . .pi interactions

Single crystal X-ray analysis of an aminoethyl mannopyranoside, namely, N-(benzyloxycarbonyl)aminoethyl-2,3,4,6-tetra-O-benzoyl-alpha-D-mannopyranoside (1), shows that the compound crystallizes in the monoclinic space group P2(1), with two molecules in the unit cell. The mannopyranoside unit adopts a distorted 4C(1) conformation. An analysis of the intermolecular interactions reveals a tandem network of N-H. . .O, C-H. . .O, and C-H. . .pi interactions responsible for stabilizing the crystal lattice.     

Morphometric image analysis of follicular lesions of the thyroid

OBJECTIVE: To determine the role of image morphometry in distinguishing various follicular lesions of the thyroid in cytologic smears. STUDY DESIGN: Archival fine needle aspiration smears of 10 cases each of follicular hyperplasia, follicular adenoma, follicular carcinoma and follicular variant of papillary carcinoma were used for the study. All cases were histopathologically proven. At least 100 random nuclei from each case were subjected to analysis with an image cytometer. Area, convex area, length, width, perimeter, convex perimeter and roundness of nuclei were measured using a 40 x objective (1 pixel = 0.446 micron). RESULTS: ANOVA showed that all the nuclear variables studied were significantly different (P < .05) in follicular hyperplasia as compared to follicular carcinoma and papillary carcinoma. All nuclear variables except roundness were also significantly different (P < .05) between follicular hyperplasia and follicular adenoma. However, between follicular adenoma, follicular carcinoma and papillary carcinoma there was considerable overlap of nuclear morphometric parameters. CONCLUSION: Image morphometry may help to distinguish nonneoplastic follicular lesions (hyperplasia) from neoplastic lesions (adenomas and carcinomas). However, to distinguish benign from malignant follicular lesions, image morphometry might not improve the accuracy of standard cytologic examination.     

Early localization of NPA58, a rat nuclear pore-associated protein, to the reforming nuclear envelope during mitosis

We have studied the mitotic reassembly of the nuclear envelope, using antibodies to nuclear marker proteins and NPA58 in F-111 rat fibroblast cells. In earlier studies we have proposed that NPA58, a 58 kDa rat nuclear protein, is involved in nuclear protein import. In this report, NPA58 is shown to be localized on the cytoplasmic face of the envelope in interphase cells, in close association with nuclear pores. In mitotic cells NPA58 is dispersed in the cytoplasm till anaphase. The targeting of NPA58 to the reforming nuclear envelope in early telophase coincides with the recruitment of a well-characterized class of nuclear pore proteins recognized by the antibody mAb 414, and occurs prior to the incorporation of lamin B1 into the envelope. Significant protein import activity is detectable only after localization of NPA58 in the newly-formed envelope. The early targeting of NPA58 is consistent with its proposed role in nuclear transport.     

Actin polymerization in response to different chemoattractants is reduced in granulocytes from chronic myeloid leukemia patients

Chronic myeloid leukemia (CML), a hematopoietic stem cell disorder, is characterized by the presence of Philadelphia chromosome (Ph1). Earlier studies have shown that various functions, such as chemotaxis, fluid phase pinocytosis, phagocytosis, and degranulation in response to chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP), were defective in polymorphonuclear leukocytes (PMNL) from CML patients. These functions depend on actin microfilaments (MF). Further studies showed that fMLP-induced actin polymerization was lower in CML PMNL. To see if this defect is specific to stimulation by fMLP alone or is a global phenomenon involving other chemoattractant receptors, chemotaxis and actin polymerization were studied in response to fMLP, an analog of fMLP, formyl-methionine-1 aminocyclooctane 1 carboxylic acid-phenyalanine-O-methionine (FACC8), platelet-activating factor (PAF), and leukotriene B4 (LTB4). These compounds bind to different chemoattractant receptors. Chemotaxis and actin polymerization in response to all four chemoattractants were significantly lower in CML PMNL compared with PMNL from normal subjects and were differentially affected for the different chemoattractants. These results suggest a global phenomenon involving all four chemoattractant-stimulated pathways. This lower amount of F-actin may be responsible for the defective chemotaxis seen in these cells.     

C-terminal region of DNA ligase IV drives XRCC4/DNA ligase IV complex to chromatin

DNA ligase IV (LIG4) and XRCC4 form a complex to ligate two DNA ends at the final step of DNA double-strand break (DSB) repair through non-homologous end-joining (NHEJ). It is not fully understood how these proteins are recruited to DSBs. We recently demonstrated radiation-induced chromatin binding of XRCC4 by biochemical fractionation using detergent Nonidet P-40. In the present study, we examined the role of LIG4 in the recruitment of XRCC4/LIG4 complex to chromatin. The chromatin binding of XRCC4 was dependent on the presence of LIG4. The mutations in two BRCT domains (W725R and W893R, respectively) of LIG4 reduced the chromatin binding of LIG4 and XRCC4. The C-terminal fragment of LIG4 (LIG4-CT) without N-terminal catalytic domains could bind to chromatin with XRCC4. LIG4-CT with W725R or W893R mutation could bind to chromatin but could not support the chromatin binding of XRCC4. The ability of C-terminal region of LIG4 to interact with chromatin might provide us with an insight into the mechanisms of DSB repair through NHEJ.     

Insulin Receptor Signaling in Cones

In humans, age-related macular degeneration and diabetic retinopathy are the most common disorders affecting cones. In retinitis pigmentosa (RP), cone cell death precedes rod cell death. Systemic administration of insulin delays the death of cones in RP mouse models lacking rods. To date there are no studies on the insulin receptor signaling in cones; however, mRNA levels of IR signaling proteins are significantly higher in cone-dominant neural retina leucine zipper (Nrl) knock-out mouse retinas compared with wild type rod-dominant retinas. We previously reported that conditional deletion of the p85α subunit of phosphoinositide 3-kinase (PI3K) in cones resulted in age-related cone degeneration, and the phenotype was not rescued by healthy rods, raising the question of why cones are not protected by the rod-derived cone survival factors. Interestingly, systemic administration of insulin has been shown to delay the death of cones in mouse models of RP lacking rods. These observations led to the hypothesis that cones may have their own endogenous neuroprotective pathway, or rod-derived cone survival factors may be signaled through cone PI3K. To test this hypothesis we generated p85α^−/−/Nrl^−/− double knock-out mice and also rhodopsin mutant mice lacking p85α and examined the effect of the p85α subunit of PI3K on cone survival. We found that the rate of cone degeneration is significantly faster in both of these models compared with respective mice with competent p85α. These studies suggest that cones may have their own endogenous PI3K-mediated neuroprotective pathway in addition to the cone viability survival signals derived from rods.     

In vitro plantlet regeneration and Agrobacterium tumefaciens-mediated genetic transformation of Indian Kino tree (Pterocarpus marsupium Roxb.)

The objective of the present study was to develop a protocol for in vitro plantlet regeneration and Agrobacterium tumefaciens-mediated genetic transformation using immature cotyledon explants of Indian Kino tree (Pterocarpus marsupium Roxb.). Immature cotyledon explants excised from 9-day-old axenic seedlings produced optimal callus on Murashige and Skoog (MS) medium supplemented with 1.07 μM α-naphthalene acetic acid (NAA), after 2 weeks of culture. When the above said callus was incubated on MS + 8.90 μM 6-benzylaminopurine (BAP) + 1.07 μM NAA, a regeneration frequency of 60.41 % with shoot number and length 12.2 ± 0.85 and 1.4 ± 0.13, respectively, was observed. For further shoot multiplication and elongation, these cultures were transferred onto MS + 4.40 μM BAP. Elongated shoots dipped in 19.60 μM indole-3-butyric acid (IBA) for 24 h and then cultured on ½MS + 2.85 μM IBA, 75 % shoots developed roots and 95 % of plantlets survived in field condition. Organogenic callus was co-cultivated with the A. tumefaciens strain LBA4404 harboring the binary plasmid pCAMBIA1301with ß-glucuronidase (uidA) and hygromycin phosphotransferase (hpt) genes and grown on MS + 8.90 μM BAP + 1.07 μM NAA (RM) + 200 μM acetosyringone for 2 days and then transferred to MS + 8.90 μM BAP + 1.07 μM NAA + 20 mg/l hygromycin + 250 mg/l cefotaxime (SIM) and 4.40 μM BAP + 15 mg/l hygromycin + 200 mg/l cefotaxime (SEM). The putatively transformed shoots were subsequently rooted on ½MS + 2.85 μM IBA + 20 mg/l hygromycin (SRM), after pulse treatment for 24 h with 19.60 μM IBA. Successful gene transfer into putatively transformed plantlets was confirmed by histochemical GUS assay, PCR and RT-PCR analysis. Southern blot analysis of regenerated plantlets confirmed the integration of hpt gene in transgenic plantlets. In the present study, a rate of 20.92 % transformation frequency was achieved and the genetic transformation protocol presented here may pave way for genetic manipulation of this multipurpose legume tree.