A quest for cytocompatible metal organic frameworks in non-viral gene therapy: Relevance of zeolitic imidazolate framework-8.

Metal-organic frameworks (MOFs) are an emerging group of nanomaterials for successful biomedical applications in gene therapy. The most commonly biocompatible MOFs are zinc-based ZIFs, zirconium-based UiOs, and iron-based MILs. However, despite increasing applications, a comparative study to underscore the critical factors for determining effective gene delivery by such MOFs is lacking. Herein, we evaluate the potential of UiO-66 and MIL-88B and ZIF-8 for gene therapeutics delivery; revealing the comparative importance of ZIF-8. Cytotoxicity assays proved insufficient for selecting the ideal gene delivery MOF vehicle. Synthesis conditions such as ability of the MOF scaffold to envelop the gene during in-situ synthesis, post-treatment such as washing, and gene loading efficiency proved to be the critical factors in determining the favourable MOF from the material selection perspective. Rapid in-situ synthesis under physiological conditions, successful gene loading, and low concentration requirements favour ZIF MOFs as gene delivery vehicles. Impact on cellular physiology, metabolism, and architecture revealed neutrality of the delivery system; and relative effects on pro-inflammatory and anti-inflammatory cytokines suggest immunomodulatory impact.

Role of the stability of charge ordering in exchange bias effect in doped manganites.

In this work we have carried out an elaborate study on the magnetic properties and investigated the exchange bias phenomena of some charge-ordered (CO) manganites. The detailed study of Sm1-x Ca x MnO3 (x = 0.5, 0.55, 0.6, 0.65, 0.7) compounds shows that Sm0.4Ca0.6MnO3, which is the most robust charge ordered material studied here, shows significantly large exchange bias field (HE) as compared to the other compounds. Our experimental results and analysis indicate that TCO, which reflects the stability of the charge-ordered state, is one of the key parameters for the exchange bias effect. Similar behaviour is found in other rare-earth analogues, viz., La1-x Ca x MnO3 and Pr1-x Ca x MnO3 compounds as well. We also found that with increasing stability of CO states in Sm1-x Ca x MnO3 compounds, HE enhances due to increase in number and reduction in size of ferromagnetic clusters.

Paraneoplastic polymyositis presenting as a clinically occult breast cancer.

Paraneoplastic syndrome affects less than 1% of cancer patients. Diagnosis of paraneoplastic syndrome with neurological presentation requires screening for an underlying malignancy, including a complete history, physical examination and imaging studies. Treatment often results in symptom stability, rather than improvement. Paraneoplastic polymyositis can precede or instantaneously occur at diagnosis or treatment of a primary tumour, while neurological symptoms can persist even following cancer treatment. We report a rare case of metaplastic breast carcinoma with an unusual presentation of paraneoplastic polymyositis.

Absence of low energy magnetic spin-fluctuations in isovalently and aliovalently doped LaCo2B2 superconducting compounds.

Magnetization, resistivity and (11)B, (59)Co NMR measurements have been performed on the Pauli paramagnet [Formula: see text], and the superconductors [Formula: see text] ([Formula: see text] K) and [Formula: see text] ([Formula: see text] K). The site selective NMR experiment reveals the multiband nature of the Fermi surface in these systems. The temperature independent Knight shift and 1/T 1 T clearly indicate the absence of correlated low energy magnetic spin-fluctuations in the normal state, which is in contrast to other Fe-based pnictides. The density of states (DOS) of Co 3d electrons has been enhanced in superconducting [Formula: see text] and [Formula: see text] with respect to the non superconducting reference compound [Formula: see text]. The occurrence of superconductivity is related to the DOS enhancement.

Significant enhancement of magnetoresistance with the reduction of particle size in nanometer scale.

The Physics of materials with large magnetoresistance (MR), defined as the percentage change of electrical resistance with the application of external magnetic field, has been an active field of research for quite some times. In addition to the fundamental interest, large MR has widespread application that includes the field of magnetic field sensor technology. New materials with large MR is interesting. However it is more appealing to vast scientific community if a method describe to achieve many fold enhancement of MR of already known materials. Our study on several manganite samples [La(1-x)Ca(x)MnO3 (x = 0.52, 0.54, 0.55)] illustrates the method of significant enhancement of MR with the reduction of the particle size in nanometer scale. Our experimentally observed results are explained by considering model consisted of a charge ordered antiferromagnetic core and a shell having short range ferromagnetic correlation between the uncompensated surface spins in nanoscale regime. The ferromagnetic fractions obtained theoretically in the nanoparticles has been shown to be in the good agreement with the experimental results. The method of several orders of magnitude improvement of the magnetoresistive property will have enormous potential for magnetic field sensor technology.

Evidence of a structural phase transition in superconducting SmFeAsO(1-x)F(x) from 19F NMR.

We report resistivity, magnetization and (19)F NMR results in a polycrystalline sample of SmFeAsO(0.86)F(0.14). The resistivity and magnetization data show a sharp drop at 48 K indicating a superconducting transition. The nuclear spin-lattice rate (1/T(1)) and spin-spin relaxation rate (1/T(2)) clearly show the existence of a structural phase transition near 163 K in the sample, which also undergoes a superconducting transition. This finding creates interest in exploring whether this is unique for Sm based systems or is also present in other rare-earth based 1111 superconductors.

Standards of care provided by early pregnancy assessment units (EPAU): a UK-wide survey.

Standards of services provided by the EPAUs across the UK vary from one unit to another. The aim of this purposive sampling self-administered survey was to assess these standards against a benchmark set by the Royal College of Obstetricians and Gynaecologists (RCOG). These standards were set out in a report by a RCOG working party (2008). Out of 181 units contacted in this survey, 140 units responded. We looked at the setup of the EPAU, services offered, accessibility and protocols for management of miscarriage and ectopic pregnancy. The survey shows that there is a considerable variation in the management protocols and the quality of services offered by the EPAUs in the UK. Many units do not meet the standards set by the RCOG.

Chaperone-like activity of tubulin. binding and reactivation of unfolded substrate enzymes.

The eukaryotic cytoskeletal protein tubulin is a heterodimer of two subunits, alpha and beta, and is a building block unit of microtubules. In a previous communication we demonstrated that tubulin possesses chaperone-like activities by preventing the stress-induced aggregation of various proteins (Guha, S., Manna, T. K., Das, K. P., and Bhattacharyya, B. (1998) J. Biol. Chem. 273, 30077-30080). As an extension of this observation, we explored whether tubulin, like other known chaperones, also protected biological activity of proteins against thermal stress or increased the yields of active proteins during refolding from a denatured state. We show here that tubulin not only prevents the thermal aggregation of alcohol dehydrogenase and malic dehydrogenase but also protects them from loss of activity. We also show that tubulin prevents the aggregation of substrates during their refolding from a denatured state and forms a stable complex with denatured substrate. The activity of malic dehydrogenase, alpha-glucosidase, and lactate dehydrogenase during their refolding from urea or guanidium hydrochloride denatured states increased significantly in presence of tubulin compared with that without tubulin. These results suggest that tubulin, in addition to its role in mitosis, cell motility, and other cellular events, might be implicated in protein folding and protection from stress.

Role of the carboxy-termini of tubulin on its chaperone-like activity.

Mutational analysis and the enzymatic digestion of many chaperones indicate the importance of both hydrophobic and hydrophilic residues for their unique property. Thus, the chaperone activity of alpha-crystallin is lost due to the substitution of hydrophobic residues or upon enzymatic digestion of the negatively charged residues. Tubulin, an eukaryotic cytoskeletal protein, exhibits chaperone-like activity as demonstrated by prevention of DTT-induced aggregation of insulin, thermal aggregation of alcohol dehydrogenase, betagamma-crystallin, and other proteins. We have shown that the tubulin lost its chaperone-like activity upon digestion of its negatively charged C-termini. In this article, the role of the C-terminus of individual subunits has been investigated. We observe that the digestion of C-terminus of beta-subunit with subtilisin causes loss of chaperone-like activity of tubulin. The contribution of C-terminus of alpha-subunit is difficult to establish directly as subtilisin cleaves C-terminus of beta-subunit first. This has been ascertained indirectly using a 14-residue peptide P2 having the sequence corresponding to a conserved region of MHC class I molecules and that binds tightly to the C-terminus of alpha-subunit. We have shown that the binding of P2 peptide to alphabeta-tubulin causes complete loss of its chaperone-like activity. NMR and gel-electrophoresis studies indicate that the P2 peptide has a significant higher binding affinity for the C-terminus of alpha-subunit compared to that of beta-subunit. Thus, we conclude that both the C-termini are necessary for the chaperone-like activity of tubulin. Implications for the chaperone functions in vivo have been discussed.

The IML3/MCM19 gene of Saccharomyces cerevisiae is required for a kinetochore-related process during chromosome segregation.

The mcm19 mutation in budding yeast affects minichromosome maintenance. In this work we have shown that this mutation leads to defects in the segregation of minichromosomes and chromosomes. The mutant cells show defective kinetochore function as judged by three criteria-- relaxation of the transcriptional block normally associated with a CEN box, stable maintenance of a dicentric plasmid in mutant cells, and mild sensitivity to the antimicrotubule drug benomyl. The MCM19 gene has been cloned and found to be the same as IML3, which codes for the ORF YBR107C. Deletion of the gene was not lethal, nor did it confer any growth defects on the mutant cells. However, the mcm19 null mutation conferred growth defects in the presence of a mutation in the TUB1 gene coding for alpha-tubulin. Two-hybrid experiments showed an interaction between Im13p/Mcm19p and the kinetochore protein Ch14, indicating that the Im13/Mcm19 protein has a role in kinetochore function.

The spindle checkpoint of the yeast Saccharomyces cerevisiae requires kinetochore function and maps to the CBF3 domain.

We have measured the activity of the spindle checkpoint in null mutants lacking kinetochore activity in the yeast Saccharomyces cerevisiae. We constructed deletion mutants for nonessential genes by one-step gene replacements. We constructed heterozygous deletions of one copy of essential genes in diploid cells and purified spores containing the deletion allele. In addition, we made gene fusions for three essential genes to target the encoded proteins for proteolysis (degron alleles). We determined that Ndc10p, Ctf13p, and Cep3p are required for checkpoint activity. In contrast, cells lacking Cbf1p, Ctf19p, Mcm21p, Slk19p, Cse4p, Mif2p, Mck1p, and Kar3p are checkpoint proficient. We conclude that the kinetochore plays a critical role in checkpoint signaling in S. cerevisiae. Spindle checkpoint activity maps to a discreet domain within the kinetochore and depends on the CBF3 protein complex.

MCM21 and MCM22, two novel genes of the yeast Saccharomyces cerevisiae are required for chromosome transmission.

The minichromosome maintenance genes, MCM21 and MCM22, have been cloned and are shown to code for the ORFs YDR318W and YJR135C respectively. Mutations in these genes caused a decrease in the stability of the minichromosome. This decrease in stability was associated with an increase in the copy number of the minichromosome in cells carrying it. Small circular dicentric plasmids were maintained relatively stably and structurally intact in the mutants compared with the wild-type strain. In the latter, such plasmids were mitotically unstable and, upon recovery, showed frequent rearrangements of their DNA. A centromere offered less obstruction to transcription in mutant cells than in the wild type, showing that both these mutants had a more relaxed kinetochore assembly. The mutant strains showed elevated rates of chromosome loss but not those of recombination. Both the mutations caused the cells to display a higher sensitivity towards the anti-mitotic drug benomyl. All these observations suggest that MCM21 and MCM22 are important for chromosome segregation with a potential role in kinetochore function. These genes are non-essential, as their deletions from chromosomes did not cause loss of cell viability. However, exponentially growing mutant cells carrying the deletion of the MCM21 gene had a significant population of large-budded cells with a single nucleus at the neck. Furthermore, the DNA content of these cells showed a shift towards 2N, suggesting a temporary pause of cells in G2 or in an early phase of mitosis. The mcm21 and mcm22 mutations do not show synthetic lethality or any further enhancement of growth defects, implying that they could be carrying out non-overlapping functions in chromosome segregation.

Generation and characterization of a polyclonal antipeptide antibody to human glycodelin.

OBJECTIVE: To develop and characterize an antiglycodelin antibody using a 15-amino acid synthetic peptide as antigen, derived from the sequence of human glycodelin. DESIGN: We have developed a chicken antiglycodelin-derived peptide antibody and have characterized the antibody with the use of endometrial and ovarian cell lines. The antibody was also tested for its ability to detect glycodelin by ELISA assay, immunocytochemistry, and by Western blot. SETTING: Various cell lines, cell culture medium, and amniotic fluid were used in the experiments. PATIENT(S): Amniotic fluid was collected from pregnant patients in their first trimester of pregnancy. INTERVENTION(S): No intervention. MAIN OUTCOME MEASURE(S): Detection of glycodelin. RESULT(S): The cell lines RL95-2 (human endometrial carcinoma cells), OVCAR-3 (human ovarian adenocarcinoma cells), HeLa (human cervical epitheloid carcinoma cells), and EM42-D (human endometrial epithelial cells) reacted with the antibody, indicating the presence of glycodelin. A specific 45-kd protein representing glycodelin was detected by Western blot in the amniotic fluid. CONCLUSION(S): Antipeptide antibodies can be successfully used to detect and quantify the presence of glycodelin in cells and fluids.

The mcm17 mutation of yeast shows a size-dependent segregational defect of a mini-chromosome.

Mini-chromosome-maintenance (mcm) mutants were described earlier as yeast mutants which could not stably maintain mini-chromosomes. Out of these, the ARS-specific class has been more extensively studied and is found to lose chromosomes and mini-chromosomes due to a defect in the initiation of DNA replication at yeast ARSs. In the present study we have identified a number of mcm mutants which show size-dependent loss of mini-chromosomes. When the size of the mini-chromosome was increased, from about 15 kb to about 60 kb, there was a dramatic increase in its mitotic stability in these mutants, but not in the ARS-specific class of mutants. One mutant, mcm17, belonging to the size-dependent class was further characterized. In this mutant, cells carried mini-chromosomes in significantly elevated copy numbers, suggesting a defect in segregation. This defect was largely suppressed in the 60-kb mini-chromosome. A non-centromeric plasmid, the TRP1ARS1 circle, was not affected in its maintenance. This mutant also displayed enhanced chromosome-III loss during mitosis over the wild-type strain, without elevating mitotic recombination. Cloning and sequencing of MCM17 has shown it to be the same as CHL4, a gene required for chromosome stability. This gene is non-essential for growth, as its disruption or deletion from the chromosome did not affect the growth-rate of cells at 23 degrees C or 37 degrees C. This work suggests that centromere-directed segregation of a chromosome in yeast is strongly influenced by its length.

Receptor induction regulates the synergistic effects of substance P with IL-1 and platelet-derived growth factor on the proliferation of bone marrow fibroblasts.

The neuropeptide substance P (SP) stimulates CFU in bone marrow (BM) cultures. Although the methylcellulose matrix used in these assays does not provide an appropriate substratum to support adherent-dependent cells, we have observed that cultures containing optimal SP (10(-8)-10(-10) M) develop confluent areas of reticular/fibroblastoid-like cells with CFUs predominantly localized within their vicinity. Characterization (cytochemical and immunofluorescence) of the reticular/fibroblastoid-like cells indicated that they were fibroblasts, the major constituent of the BM stroma. Hemopoietic effects by SP are mediated by the stroma that expresses SP receptors. We studied the effects of SP (10(-7)-10(-11) M) with suboptimal platelet-derived growth factor-BB (PDGF-BB; 5 ng/ml) and IL-1alpha (2 ng/ml), two fibrogenic cytokines, and also hemopoietic regulators. SP by itself and in synergy with either cytokine induced fibroblast proliferation. At optimum SP, IL-1alpha induced 1.6 times the proliferation of PDGF-BB (87 +/- 7 vs 55 +/- 5; n = 12; p < 0.05). The effects of SP were blunted by a specific neurokinin-1 antagonist. Scatchard analysis indicated that SP binds to BM fibroblasts with an approximate Kd of 5 nM. SP induced steady state mRNA for IL-1 receptor IL-1RI and PDGF-BB (PDGF-AR, PDGF-BR) receptors by 7.5-, 6.2-, and 10.5-fold, respectively. Their up-regulation may be partly responsible for the synergistic effects of SP and their ligands. Induction (3-fold) of neurokinin-1 mRNA by IL-1alpha compared with no induction by PDGF-BB may explain the preferred synergism between SP and IL-1alpha. This study indicates that induction of SP, IL-1alpha, and PDGF-BB receptors is important to their synergistic effects on BM fibroblast proliferation. These results bring new insights into stroma-mediated hemopoietic regulation.

Hematopoietic regulation mediated by interactions among the neurokinins and cytokines.

This review summarizes the current data regarding the mechanisms by which two mammalian neurokinins (tachykinins), substance P (SP) and neurokinin-A (NK-A) are involved in hematopoiesis. SP and NK-A are derived from the preprotachykinin-I (PPT-I) gene which can be induced by cytokines and neurotrophic factors. In the bone marrow (BM), nerve fibers and stroma are potential sources for the PPT-I gene products. SP and NK-A interact with either of three cloned receptors, neurokinin-1 (NK-1), NK-2 or NK-3, although SP and NK-A exhibit binding preferences for NK-1 and NK-2 respectively. Through specific receptors, SP and NK-A exert dichotomous hematopoietic effects mediated mostly by the BM stroma. SP enhances the proliferation of primitive BM stem cells and progenitors and these effects correlate with the induction of stimulatory hematopoietic growth factors. NK-A appears to be protective to stem cells through the induction of TGF-beta. Proliferation of myeloid progenitors is inhibited by NK-A, effects which correlate with the induction of two suppressive factors, TGF-beta and MIP-1alpha. Stimulation of NK-2 leads to partial blunting of the enhanced stimulatory effects mediated by NK-1. Furthermore, stimulatory hematopoietic cytokines upregulate NK-1 expression and downregulate the constitutively expressed NK-2 in BM stroma. Together, the experimental evidence suggests that NK-A-NK-2 interactions could be a feedback to hematopoietic stimulation. Expression of NK-1 and NK-2 in CD34+ cell lines and also, the presence of SP binding sites on primary CD34+ cells suggest that the neurokinins could be interacting directly with BM progenitors and stem cells. In BM stroma, cytokines and neurokinins regulate the expression of each other and also, their respective receptors. In summary, the current literature pertaining to hematopoietic regulation indicates the involvement of a complex network that includes, but not exclusive of the cytokines and neurokinins. The current models that pertain to stem cell proliferation and differentiation should therefore add neuropeptides to the list of hematopoietic modulators.

Perception about AIDS among residents of a Calcutta slum.

A study was carried out in a slum area of South Calcutta to assess the impact of the current mass education programme against AIDS. Two hundred and six residents, mainly of lower middle class, aged 18-60 years of both sexes were selected at random. They were interviewed to know their perception and sources of information about AIDS. Two-third of them had their own TV and radio, which they watched/heard for about three and half hours each day. Another 28% watched TV outside for about one and half hours a day. About 46% were daily readers and 20% occasional readers of newspapers. Fifty nine percent knew about persons vulnerable to get AIDS, but most of them associated it to promiscuity only. Avoiding it was the main means known to them for preventing AIDS. The role of condom in it was known to only 2.5% residents. The source of their knowledge was mainly TV, either alone or with other mass media (67%). Such knowledge was related to higher education. To make perception about AIDS more effective, it is suggested that local health and voluntary agencies should involve the community in the AIDS education programme along with the back-up of mass media. An apex agency solely responsible for AIDS education should be set up for each big city to co-ordinate the activities of local agencies.

PIP: 206 mainly poor and lower middle-class residents of the Chetla slum of South Calcutta were interviewed with regard to their perceptions and sources of information about AIDS. The goal was to assess the impact of the current mass education program against AIDS. One member was randomly selected from each of 206 families to participate in the study. Respondents were aged 16-80 years, of mean age 35-42 years, 43.7% male, 80% married, 19% illiterate, and 94% Hindu. 66% had their own television and radio which they watched/heard for approximately 3.5 hours daily. Another 28% watched television outside for approximately 1.5 hours each day. Approximately 46% and 20% were daily and occasional newspaper readers, respectively. 59% knew about people vulnerable to contracting AIDS, but most associated HIV risk only with promiscuity and prostitution. Avoiding such behavior was deemed to be the best way to avoid contracting HIV. Only 2.5% knew that condom use can protect against HIV infection. The source of that knowledge was mainly television, either alone or with other mass media. Such knowledge was related to higher education. The authors suggest that local health and voluntary agencies involve the community in the AIDS education program, backed up by the mass media. An agency solely responsible for AIDS education should be established in each major city to coordinate local agency activities.

Predatory aggression induced by hypothalamic stimulation: modulation by midbrain periaqueductal gray (PAG).

Adequate electrical stimulation of extreme lateral hypothalamic regions of healthy, non-aggressive male cats was employed to produce aggression on live but anaesthetized rats. Stimulus response (S-R) curves based on scoring systems for both somatic and affective display components of behaviour were used to assess how manipulation of midbrain PAG by electrocoagulative lesions or drug microinjections affected the sensitivity of attack producing hypothalamic loci. Anodal lesions of dorsal PAG and adjoining tectum increased the excitability of hypothalamic loci producing predatory attack. Microinjection of 250 ng of delta-alanine-methionine enkephalin (DAME) in dPAG completely suppressed the somatomotor components of attack behaviour and markedly inhibited the affective display components. Administration of naloxone, an opioid antagonist (1 microgram) at the same sites facilitated the hypothalamically induced attack behaviour and annulled the inhibitory effect of DAME. These findings indicate the involvement of midbrain enkephalinergic mechanisms in the modulation of predatory attack behaviour elaborated by hypothalamic stimulation.

Fine needle aspiration cytology of cervical lymphadenopathy with special reference to tuberculosis.

One hundred eighty cases of cervical lymphadenopathy have been studied by fine needle aspiration cytological examination followed by histopathologic examination of the excised lymph nodes. The diagnostic accuracy was 84.4% for tuberculous lymphadenitis by fine needle aspiration cytological examination. Observation of caseous necrosis (84.2%) and epithelioid cells (73.6%) were the most characteristic diagnostic features in the aspirated smears. Acid-fast bacilli were observed in 45.6% cases. Metastatic carcinoma also yielded a high diagnostic accuracy ie, 89%. Fine needle aspiration cytology has been found to be safe, quick, inexpensive with high diagnostic accuracy in cervical lymphadenopathy.