Submucosal Diclofenac for Acute Postoperative Pain in Third Molar Surgery: A Randomized, Controlled Clinical Trial.

Diclofenac sodium is a widely used nonsteroidal anti-inflammatory drug (NSAID) for relief of inflammatory pain. A recent formulation combines this drug with hydroxypropyl-ß-cyclodextrin (HPßCD) to improve its solubility and to enable subcutaneous administration. Previous studies confirmed the efficacy of this combination. This study's aim was to evaluate the efficacy, safety, and local tolerability of diclofenac HPßCD administered as a local submucosal injection prior to lower third molar surgery. We conducted a prospective, randomized, double-blind, placebo-controlled, parallel-group phase II single-center study. Seventy-five patients requiring mandibular third molar surgery were randomized into 1 of 5 groups: 5 mg/1 mL diclofenac HPßCD, 12.5 mg/1 mL diclofenac HPßCD, 25 mg/1 mL diclofenac HPßCD, 50 mg/1 mL diclofenac HPßCD, or 1 mL placebo. The respective study drug was injected into the mucosal tissue surrounding the surgical site prior to surgery following achievement of local anesthesia. The primary outcome measure was the area under the curve (AUC) of cumulative pain scores from end of surgery to 6 h postsurgery. This demonstrated a global treatment effect between the active groups and placebo, hence confirming the study drug's efficacy ( P = 0.0126). Secondary outcome measures included the time until onset of pain and the time until patients required rescue medication, both showing statistical significance of the study drug compared to placebo ( P < 0.0161 and P < 0.0001, respectively). The time until rescue medication ranged between 7.8 h (for 25 mg/1 mL diclofenac HPßCD) and 16 h (for 50 mg/1 mL diclofenac HPßCD). Interestingly, the 5-mg/1-mL solution appeared superior to the 12.5-mg/1-mL and 25-mg/1-mL solutions (time until rescue medication = 12.44 h). A total of 14% of patients experienced minor adverse drug reactions (ADRs), of which 2 cases demonstrated flap necrosis. These resolved without further intervention. The study results overall indicate efficacy, safety, and relative tolerability of diclofenac HPßCD used locally as a submucosal injection prior to third molar surgery (ClinicalTrials.gov NCT01706588).

Prognostic value of the hDMP1-ARF-Hdm2-p53 pathway in breast cancer.

Our recent study showed critical roles of Dmp1 as a sensor of oncogenic Ras, HER2/neu signaling and activation of the Arf-p53 pathway. To elucidate the role of human DMP1 (hDMP1) in breast cancer, one hundred and ten pairs of human breast cancer specimen were studied for the alterations of the hDMP1-ARF-Hdm2-p53 pathway with follow up of clinical outcomes. Loss of heterozygosity (LOH) of the hDMP1 locus was found in 42% of human breast carcinomas, while that of INK4a/ARF and p53 were found in 20 and 34%, respectively. Hdm2 amplification was found in 13% of the same sample, which was found independently of LOH for hDMP1. Conversely, LOH for hDMP1 was found in mutually exclusive fashion with that of INK4a/ARF and p53, and was associated with low Ki67 index and diploid karyotype. Consistently, LOH for hDMP1 was associated with luminal A category and longer relapse-free survival, while that of p53 was associated with non-luminal A and shorter survival. Thus, loss of hDMP1 could define a new disease category associated with prognosis of breast cancer patients. Human breast epithelial cells/cancer cells with wild-type p53 were sensitive to growth inhibition by activated Dmp1:ER while those that delete p14(ARF) or p53, and/or Hdm2 amplification showed partial or nearly complete resistance, indicating that p53 is a critical target for hDMP1 to exhibit its biological activity.

Repression of Dmp1 and Arf transcription by anthracyclins: critical roles of the NF-kappaB subunit p65.

Both genotoxic and oncogenic stress activates the nuclear factor-kappa B (NF-kappaB) and p53 proteins; however, the p53 activity is antagonized by NF-kappaB signaling. Dmp1 is a Myb-like transcription factor that activates the Arf-p53 pathway. The Dmp1 promoter was activated by a classical NF-kappaB activator tumor necrosis factor alpha, but repressed by treatment of cells with non-classical NF-kappaB activators, anthracyclins and UV-C. p65 and other subsets of NF-kappaB proteins were bound to the Dmp1 promoter following anthracyclin/UV-C treatment of rodent fibroblasts. This resulted in the downregulation of Dmp1 mRNA and protein. Repression of the Dmp1 transcription by anthracyclins depended on the unique NF-kappaB site on the promoter. Downregulation of p65 significantly attenuated the repression of the Dmp1 promoter by anthracyclins/UV-C. The amount of Dmp1 bound to the Arf promoter decreased significantly upon anthracyclin treatment; this, in turn, downregulated the Arf levels. Repression of the Arf promoter by p65 or anthracyclins depended on Dmp1, which was significantly attenuated in Dmp1(-/-) cells. Both Dmp1(-/-)and Arf(-/-)cells showed resistance to anthracyclin-induced cell death compared to wild-type cells; non-immortalized p65-knockdown cells were much more sensitive. Thus, the Dmp1-Arf pathway is repressed by p65 in response to genotoxic stress, which implicates a novel mechanism of p53 inactivation by NF-kappaB.

Expression of Dmp1 in specific differentiated, nonproliferating cells and its regulation by E2Fs.

Dmp1 is a Myb-like transcription factor that transmits oncogenic Ras-Raf signaling to the Arf-p53 pathway and induces cell cycle arrest. Immunohistochemical staining was performed to identify the pattern of Dmp1 expression in normal murine tissues compared with the proliferation marker, Ki67. In thymus, the nuclei of mature T lymphocytes in the medulla were strongly positive for Dmp1, whereas Ki67 was detected only in the cortex. In intestine, Dmp1 was detected in the nuclei of superficial layers of the villi, whereas Ki67-positive cells were confined to the lower one-third of the crypt. Double staining for Dmp1 and Ki67 revealed that these two proteins were expressed in mutually exclusive fashion in nearly all the tissues examined. Subsets of E2Fs were specifically bound to the Dmp1 promoter upon mitogenic signaling and E2Fs 1-4 inhibited the Dmp1 promoter in a reporter assay. The Dmp1 promoter was repressed when the cells entered the S to G2/M phase of the cell cycle when both Dmp1 and Arf expressions were downregulated. The Dmp1 mRNA was not downregulated by serum in E2F-DB(+) cells, suggesting that the Dmp1 promoter repression is E2F-dependent. This explains why the Dmp1 and Ki67-positive cells are stained in mutually exclusive fashion in normal tissues.

Metal nanoplasmas as bright sources of hard X-ray pulses.

We report significant enhancements in light coupling to intense-laser-created solid plasmas via surface plasmon and "lightning rod" effects. We demonstrate this in metal nanoparticle-coated solid targets irradiated with 100 fs, 806 nm laser pulses, focused to intensities approximately 10(14)-10(15) W cm(-2). Our experiments show a 13-fold enhancement in hard x-ray yield (10-200 keV) emitted by copper nanoparticle plasmas formed at the focal volume. A simple model explains the observed enhancement quantitatively and provides pointers to the design of structured surfaces for maximizing such emissions.

Antimutagenic effect of black tea extract using 'rodent dominant lethal mutation assay'.

The antimutagenic effect of black tea extract has been evaluated with the 'Dominant Lethal Assay' in Swiss albino mice using benzo[a]pyrene [BaP] as a mutagen. BaP was given through the intraperitoneal (i.p.) route at a single dose of 100 mg/kg b.w. to male mice once only. The animals were given 1, 2 and 4% aqueous solution of black tea as sole source of drinking solution prior to BaP. The pregnant females were analyzed for living implants, pre- and post-implantation losses. The results revealed that during mating weeks, BaP caused a reduction in implants and an increase in pre- and post-implantation losses. The protective effect of tea solution on BaP-induced mutagenicity was observed. The number of living implants increased and dead implants decreased significantly in the animals kept on 2 and 4% tea solution. The increase in dominant lethal mutation rate by BaP was inhibited by black tea extract. Four percent tea solution alone did not produce dominant lethality, and reveals that it is non-toxic/non-mutagenic to sperm. Hence the study suggests that tea has a protective effect against BaP-induced genetic damage to germ cells in Swiss albino mice.

The liver in choledochal cyst.

BACKGROUND/PURPOSE: Hepatic changes due to choledochal cyst have not been given due emphasis in the published literature. In this study the gross and microscopic appearance of the liver in patients with choledochal cyst have been correlated with clinical features, liver function tests and postoperative complications. METHODS: A retrospective evaluation of patients treated for choledochal cysts between January 1989-December 1998 was undertaken. The case records were reviewed and liver biopsies taken at surgery were analysed. RESULTS: The case records of 22 patients were reviewed (16 girls, 6 boys; mean age 4.6 years, range 1 month-10 years). The presenting features included jaundice (59%; 85.7% in < 1 year), abdominal pain (59%; 86.7% in > 1 years) and fever in 40.9% cases. A palpable abdominal mass and hepatomegaly was present in 32% cases. A type IV cyst was present in 1 case; all others had type I cysts (14 cystic and 7 fusiform). The liver looked grossly 'normal' in 16 and 'cirrhotic' in 6 cases. Liver biopsies were available for review from 5 of the 'cirrhotic' and 7 of the 'normal' looking livers. All the liver biopsies showed varying degrees of bile duct proliferation, cholestasis, parenchymal damage, inflammatory cell infiltration and pericentral fibrosis. Histological features of cirrhosis were evident in 6 cases (4 'cirrhotic' and 2 'normal' looking livers), 4 of these cases were infants. Liver function tests were deranged in 5 cases with histological features of cirrhosis and hepato-biliary scintigraphy showed obstructive features in 3 of these cases. There was an increased risk of postoperative complications in the cases with histological features of cirrhosis, 2 died from hepatic insufficiency and one each had transient ascitic and biliary leak. CONCLUSION: Liver histology showed significant changes in all the cases of choledochal cyst in whom it was studied; even normal looking livers showed evidence of significant changes. Presence of cirrhosis, more common in infants, correlated with jaundice, deranged liver function tests, obstructive features on hepatobiliary scintigraphy and a greater risk of postoperative complications.

Villoglandular papillary adenocarcinoma of the cervix: case report.

The incidence of cervical adenocarcinoma is on the rise over the last several decades. It generally carries a worse prognosis than squamous carcinoma. Villoglandular papillary adenocarcinoma (VGA) of the cervix has been identified as a distinct subset of cervical adenocarcinoma that occurs in young women and supposedly carries an excellent prognosis. We report a case of adenocarcinoma of the cervix in a young woman that had classical histological features of VGA but at operation showed presence of tumor cells in the peritoneal wash. A review of world literature on the 56 cases reported so far is presented where occasional cases showing disease spread have been reported, suggesting caution in the management and follow-up of this clinicopathologic entity.

Primary malignant bladder carcinoid--a diagnostic and management dilemma.

A middle-aged man who presented with gross hematuria and irritative voiding symptoms was found to have an invasive infiltrating primary bladder carcinoid after radical cystoprostatectomy. Hurricane chest secondaries were prominent in the follow-up evaluation. The immunohistochemical markers for neuroendocrine differentiation were substantiated. The published reports of primary bladder carcinoid tumors are discussed.

Mutagenic evaluation of deltamethrin using rodent dominant lethal assay.

The dominant lethal test was used to analyse the mutagenic potential of deltamethrin, a synthetic pyrethroid insecticide, in Swiss albino mice. In the treated series, the animals were exposed orally to three different doses (0.36, 0.72 and 1.08 mg/kg body weight) of deltamethrin dissolved in corn oil. Following the treatment, each male of control, as well as of the treated series, was mated with untreated females, every week for a period of 6 weeks. All mated females were sacrificed on the 13th day of separation and their ovaries and uterus were examined. The results revealed that deltamethrin treatment did not impair the mating capacity and fertility of Swiss albino mice. Mutagenic index, pre- and post-implantation losses were assessed. No significant pre-implantation losses were observed either weekly or averagely. Post-implantation losses were observed at medium and high doses of deltamethrin. A slight increase in dominant lethal mutation rate was observed by increasing doses of deltamethrin in early weeks but decreased in later weeks, so an apparent dose response was not observed.

Constituents of the yew trees.

Yew trees, taxonomically classified under the genus Taxus, are sources of a number of physiologically active compounds of different classes. Taxane derivatives with various carbon skeletons, lignans, flavonoids, steroids and sugar derivatives have been isolated from different Taxus species. Compounds isolated from the genus Taxus between 1908 and December 1997 have been comprehensively reviewed.

Two regions of simian virus 40 T antigen determine cooperativity of double-hexamer assembly on the viral origin of DNA replication and promote hexamer interactions during bidirectional origin DNA unwinding.

Phosphorylation of simian virus 40 large tumor (T) antigen on threonine 124 is essential for viral DNA replication. A mutant T antigen (T124A), in which this threonine was replaced by alanine, has helicase activity, assembles double hexamers on viral-origin DNA, and locally distorts the origin DNA structure, but it cannot catalyze origin DNA unwinding. A class of T-antigen mutants with single-amino-acid substitutions in the DNA binding domain (class 4) has remarkably similar properties, although these proteins are phosphorylated on threonine 124, as we show here. By comparing the DNA binding properties of the T124A and class 4 mutant proteins with those of the wild type, we demonstrate that mutant double hexamers bind to viral origin DNA with reduced cooperativity. We report that T124A T-antigen subunits impair the ability of double hexamers containing the wild-type protein to unwind viral origin DNA, suggesting that interactions between hexamers are also required for unwinding. Moreover, the T124A and class 4 mutant T antigens display dominant-negative inhibition of the viral DNA replication activity of the wild-type protein. We propose that interactions between hexamers, mediated through the DNA binding domain and the N-terminal phosphorylated region of T antigen, play a role in double-hexamer assembly and origin DNA unwinding. We speculate that one surface of the DNA binding domain in each subunit of one hexamer may form a docking site that can interact with each subunit in the other hexamer, either directly with the N-terminal phosphorylated region or with another region that is regulated by phosphorylation.

Interaction of the transcription factor TFIID with simian virus 40 (SV40) large T antigen interferes with replication of SV40 DNA in vitro.

Simian virus 40 (SV40) large tumor (T) antigen is the major regulatory protein that directs the course of viral infection, primarily by interacting with host cell proteins and modulating their functions. Initiation of viral DNA replication requires specific interactions of T antigen bound to the viral origin of DNA replication with cellular replication proteins. Transcription factors are thought to stimulate initiation of viral DNA replication, but the mechanism of stimulation is poorly understood. Since the transcription factor TATA-binding protein (TBP) binds to sequences within the origin of replication and interacts specifically with T antigen, we examined whether TBP complexes stimulate SV40 DNA replication in vitro. On the contrary, we found that depletion of TBP complexes from human cell extracts increased their ability to support viral DNA replication, and readdition of TBP complexes to the depleted extracts diminished their activity. We have mapped the sites of interaction between the proteins to residues 181 to 205 of T antigen and 184 to 220 of TBP. Titration of fusion proteins containing either of these peptides into undepleted cell extracts stimulated their replication activity, suggesting that they prevented the T antigen-TBP interaction that interfered with replication activity. TBP complexes also interfered with origin DNA unwinding by purified T antigen, and addition of either the T antigen or the TBP fusion peptide relieved the inhibition. These results suggest that TBP complexes associate with a T-antigen surface that is also required for origin DNA unwinding and viral DNA replication. We speculate that competition among cellular proteins for T antigen may play a role in regulating the course of viral infection.

The replication protein A binding site in simian virus 40 (SV40) T antigen and its role in the initial steps of SV40 DNA replication.

Physical interactions of simian virus 40 (SV40) large tumor (T) antigen with cellular DNA polymerase alpha-primase (Pol/Prim) and replication protein A (RPA) appear to be responsible for multiple functional interactions among these proteins that are required for initiation of viral DNA replication at the origin, as well as during lagging-strand synthesis. In this study, we mapped an RPA binding site in T antigen (residues 164 to 249) that is embedded within the DNA binding domain of T antigen. Two monoclonal antibodies whose epitopes map within this region specifically interfered with RPA binding to T antigen but did not affect T-antigen binding to origin DNA or Pol/Prim, ATPase, or DNA helicase activity and had only a modest effect on origin DNA unwinding, suggesting that they could be used to test the functional importance of this RPA binding site in the initiation of viral DNA replication. To rule out a possible effect of these antibodies on origin DNA unwinding, we used a two-step initiation reaction in which an underwound template was first generated in the absence of primer synthesis. In the second step, primer synthesis was monitored with or without the antibodies. Alternatively, an underwound primed template was formed in the first step, and primer elongation was tested with or without antibodies in the second step. The results show that the antibodies specifically inhibited both primer synthesis and primer elongation, demonstrating that this RPA binding site in T antigen plays an essential role in both events.

Infant and child mortality in Bhil tribe of Jhabua district.

Nearly 25 million children are born in India every year of which almost 2.7 million die before attaining the age of five years. Forty seven per cent of births take place in the four states, namely Uttar Pradesh, Madhya Pradesh, Bihar and Orissa, while fifty per cent of all deaths below five years also take place in these states. The present study was carried out in Jhabua district in which five per cent villages of each tehsil were selected by random sampling. Information was obtained on 430 households of 67 villages on infant and child mortality, birth order, age, sex and cause of death. 38.2% households reported the death of at least one child below the age of five years. Out of these 59.3%, 27.2% and 13.2% reported the death of one, two or three and more children respectively. 46% of infant deaths or 29.3% of all deaths occurred in the neonatal period. The major causes of death were preventable such as tetanus, diarrhoea, measles, ARI and fever. 51.3% deaths were of children who were third or more in birth order. 54.8% deaths were males and 47.59% were females. The study shows the need for an effective Dai (Midwife) Training Programme to upgrade the skills and an urgent necessity to improve the outreach services in remote tribal areas to bring down the infant mortality.

Purification and properties of human DNA helicase VI.

A novel ATP-dependent DNA unwinding enzyme, called human DNA helicase VI (HDH VI), was purified to apparent homogeneity from HeLa cells and characterized. From 327 g of cultured cells, 0.44 mg of pure enzyme was recovered, free of DNA polymerase, ligase, topoisomerase, nicking and nuclease activities. The enzyme behaves as a monomer having an M(r) of 128 kDa, whether determined with SDS-PAGE, or in native conditions. Photoaffinity labelling with [alpha-32P]ATP labelled the 128 kDa protein. Only ATP or dATP hydrolysis supports the unwinding activity for which a divalent cation (Mg2+ > Mn2+) is required. HDH VI unwinds exclusively DNA duplexes with an annealed portion < 32 bp and prefers a replication fork-like structure of the substrate. It cannot unwind blunt-end duplexes and is inactive also on DNA-RNA or RNA-RNA hybrids. HDH VI unwinds DNA unidirectionally by moving in the 3' to 5' direction along the bound strand.

Human DNA helicase II: a novel DNA unwinding enzyme identified as the Ku autoantigen.

Human DNA helicase II (HDH II) is a novel ATP-dependent DNA unwinding enzyme, purified to apparent homogeneity from HeLa cells, which (i) unwinds exclusively DNA duplexes, (ii) prefers partially unwound substrates and (iii) proceeds in the 3' to 5' direction on the bound strand. HDH II is a heterodimer of 72 and 87 kDa polypeptides. It shows single-stranded DNA-dependent ATPase activity, as well as double-stranded DNA binding capacity. All these activities comigrate in gel filtration and glycerol gradients, giving a sedimentation coefficient of 7.4S and a Stokes radius of approximately 46 A, corresponding to a native molecular weight of 158 kDa. The antibodies raised in rabbit against either polypeptide can remove from the solution all the activities of HDH II. Photoaffinity labelling with [alpha-32P]ATP labelled both polypeptides. Microsequencing of the separate polypeptides of HDH II and cross-reaction with specific antibodies showed that this enzyme is identical to Ku, an autoantigen recognized by the sera of scleroderma and lupus erythematosus patients, which binds specifically to duplex DNA ends and is regulator of a DNA-dependent protein kinase. Recombinant HDH II/Ku protein expressed in and purified from Escherichia coli cells showed DNA binding and helicase activities indistinguishable from those of the isolated protein. The exclusively nuclear location of HDH II/Ku antigen, its highly specific affinity for double-stranded DNA, its abundance and its newly demonstrated ability to unwind exclusively DNA duplexes, point to an additional, if still unclear, role for this molecule in DNA metabolism.

Anti-invasive activity of 3,7-dimethoxyflavone in vitro.

Invasion of MCF-7/6 human mammary carcinoma cells into embryonic chick heart fragments was studied in organ culture during 8 days. The effect of 31 polyphenolic compounds, belonging to the flavonoids, chalcones, or coumarins, was tested in this assay for invasion. The anti-invasive activity of 3,7-dimethoxyflavone was found at concentrations ranging from 1 to 100 microM. At these anti-invasive concentrations, no cytotoxic effects could be detected: the anti-invasive effect was reversible upon omission of the molecule from the medium, and treatment of MCF-7/6 cells or heart fragments did not affect subsequent outgrowth from explants on tissue culture plastic. The molecule did not inhibit growth of MCF-7/6 cell aggregates nor of heart fragments kept in suspension culture. The action mechanism of 3,7-dimethoxyflavone is the subject of our ongoing research.