New Jersey Space Grant Consortium

We have determined the complete primary structure for human matrix metalloproteinase-3 (MMP-3), which has 477 residues including a 17-residue signal peptide. The result indicates that MMP-3 is identical with stromelysin (Whitham, S. E., Murphy, G., Angel, P., Rahmsdorf, H.-J., Smith, B. J., Lyons, A., Harris, T. J. R., Reynolds, J. J., Herrlich, P., and Docherty, A. J. P. (1986) Biochem. J. 240, 913-916). A striking result is that MMP-3 and collagenase are 54% identical in sequence, suggesting a common origin for the evolution of the two proteinases. We also show that in human synovial fibroblast cultures human recombinant interleukin-1 beta rapidly induces high levels of MMP-3 mRNA and, conversely, that retinoic acid or dexamethasone can suppress the MMP-3 mRNA levels. Similar results were obtained for human synovial collagenase mRNA. The data suggest that MMP-3 and collagenase expression are coordinately modulated in synovial fibroblast cultures.

Patient non-adherence to medication is a pervasive problem that contributes to poor patient health and high healthcare costs. Basic research and interventions have focused thus far on behaviour initiation factors, such as patients' illness and treatment beliefs. This paper proposes two processes that occur after behaviour initiation that are theorised to contribute to prediction of long-term medication adherence: 'coherence' of patients' beliefs from experiences with treatment and habit development. Seventy-one hypertensive patients reported their treatment-related beliefs, experiences related to treatment efficacy and medication-taking habit strength in a baseline interview. Patients then used an electronic monitoring pill bottle for approximately one month. Patients' medication habit-strength was the strongest predictor of all adherence measures, explaining 6-27% incremental variance in adherence to that explained by patients' treatment-related beliefs. Patients' beliefs and experiences did not predict overall adherence, even for patients with 'weaker' habits. However, patients' experiences were found to predict intentional non-adherence and habit strength was found to predict unintentional adherence. Practitioners may assess patients' medication-taking habits to get an initial view of their likely adherence to long-term medications. Future research should assess the current theoretical predictions in a hypertension inception sample and in populations with symptomatic conditions.

Glycan structures on glycoproteins and glycolipids play critical roles in biological recognition, targeting, and modulation of functions in animal systems. Many classes of glycan structures are capped with terminal sialic acid residues, which contribute to biological functions by either forming or masking glycan recognition sites on the cell surface or secreted glycoconjugates. Sialylated glycans are synthesized in mammals by a single conserved family of sialyltransferases that have diverse linkage and acceptor specificities. We examined the enzymatic basis for glycan sialylation in animal systems by determining the crystal structures of rat ST6GAL1, an enzyme that creates terminal α2,6-sialic acid linkages on complex-type N-glycans, at 2.4 Å resolution. Crystals were obtained from enzyme preparations generated in mammalian cells. The resulting structure revealed an overall protein fold broadly resembling the previously determined structure of pig ST3GAL1, including a CMP-sialic acid-binding site assembled from conserved sialylmotif sequence elements. Significant differences in structure and disulfide bonding patterns were found outside the sialylmotif sequences, including differences in residues predicted to interact with the glycan acceptor. Computational substrate docking and molecular dynamics simulations were performed to predict and evaluate the CMP-sialic acid donor and glycan acceptor interactions, and the results were compared with kinetic analysis of active site mutants. Comparisons of the structure with pig ST3GAL1 and a bacterial sialyltransferase revealed a similar positioning of donor, acceptor, and catalytic residues that provide a common structural framework for catalysis by the mammalian and bacterial sialyltransferases.

Developmental abnormalities of myelination are observed in the brains of laminin-deficient humans and mice. The mechanisms by which these defects occur remain unknown. It has been proposed that, given their central role in mediating extracellular matrix (ECM) interactions, integrin receptors are likely to be involved. However, it is a non-integrin ECM receptor, dystroglycan, that provides the key linkage between the dystrophin-glycoprotein complex (DGC) and laminin in skeletal muscle basal lamina, such that disruption of this bridge results in muscular dystrophy. In addition, the loss of dystroglycan from Schwann cells causes myelin instability and disorganization of the nodes of Ranvier. To date, it is unknown whether dystroglycan plays a role during central nervous system (CNS) myelination. Here, we report that the myelinating glia of the CNS, oligodendrocytes, express and use dystroglycan receptors to regulate myelin formation. In the absence of normal dystroglycan expression, primary oligodendrocytes showed substantial deficits in their ability to differentiate and to produce normal levels of myelin-specific proteins. After blocking the function of dystroglycan receptors, oligodendrocytes failed both to produce complex myelin membrane sheets and to initiate myelinating segments when co-cultured with dorsal root ganglion neurons. By contrast, enhanced oligodendrocyte survival in response to the ECM, in conjunction with growth factors, was dependent on interactions with beta-1 integrins and did not require dystroglycan. Together, these results indicate that laminins are likely to regulate CNS myelination by interacting with both integrin receptors and dystroglycan receptors, and that oligodendrocyte dystroglycan receptors may have a specific role in regulating terminal stages of myelination, such as myelin membrane production, growth, or stability.

BACKGROUND: Birthweight for gestational age is lower in US Black infants than in US White infants. It is unknown, however, whether this difference is 'normal' (i.e. physiological) or reflects pathological foetal growth restriction. METHODS: We applied an analytic approach based on foetuses at risk to compare gestational age-specific rates of live birth, 'revealed' small-for-gestational-age (SGA), and neonatal mortality among singleton infants >or=22 weeks of gestation and >or=500 g born in 1998-2000 to US White (n = 9 012 194), US-born Black (n = 1 554 382), and foreign-born Black (n = 200 395) mothers. Graphical methods and Cox proportional hazards regression analyses were used to compare outcomes in the three ethnic groups. RESULTS: Rates of live birth and neonatal mortality were highest at all gestational ages in US-born Blacks, lowest in Whites, and intermediate in foreign-born Blacks. The revealed SGA pattern cohered much more closely with the observed pattern for neonatal mortality when SGA was defined based on a single, overall standard of birthweight for gestational age than when based on ethnic group-specific standards. CONCLUSION: The closer coherence of revealed SGA and neonatal mortality rates based on a single standard and the intermediate pattern among foreign-born Blacks strongly suggest that Black-White differences in birthweight for gestational age are pathological, rather than physiological.

RNA polymerases I, II, and III each use the TATA-binding protein (TBP). Regulators that target this shared factor may therefore provide a means to coordinate the activities of the three nuclear RNA polymerases. The repressor Dr1 binds to TBP and blocks the interaction of TBP with polymerase II- and polymerase III-specific factors. This enables Dr1 to coordinately regulate transcription by RNA polymerases II and III. Under the same conditions, Dr1 does not inhibit polymerase I transcription. By selectively repressing polymerases II and III, Dr1 may shift the physiological balance of transcriptional output in favor of polymerase I.

Genetic susceptibility to multiple sclerosis (MS) has been linked to the HLA-DR15 haplotype consisting of DRB1*15:01(DR2b) and DRB5*01:01(DR2a) alleles. Given almost complete linkage disequilibrium of the two alleles, recent studies suggested differential roles in susceptibility (DR2b) or protection from MS (DR2a). Our objective was to assess the potential contribution of DR2a to disease etiology in MS using a humanized model of autoimmunity. To assess the potential contribution of DR2a to disease etiology, we created DR2a humanized transgenic (Tg) mice and subsequently crossed them to Tg mice expressing TL3A6, an MS patient-derived myelin basic protein 83-99-specific TCR. In TL3A6/DR2a Tg mice, CD4 Tg T cells escape thymic and peripheral deletion and initiate spontaneous experimental autoimmune encephalomyelitis (EAE) at low rates, depending on the level of DR2a expression. The ability to induce active EAE was also increased in animals expressing higher levels of DR2a. Inflammatory infiltrates and neuronal damage were present throughout the spinal cord, consistent with a classical ascending EAE phenotype with minor involvement of the cerebellum, brainstem, and peripheral nerve roots in spontaneous, as well as actively induced, disease. These studies emphasize the pathologic contribution of the DR2a allele to the development of autoimmunity when expressed as the sole MHC class II molecule, as well as strongly argue for DR2a as a contributor to the CNS autoimmunity in MS.

Parathyroid hormone (PTH) regulates the transcription of many genes involved in bone remodeling in osteoblasts. One of these genes is matrix metalloproteinase-13 (MMP-13), which is involved in bone remodeling and early stages of endochondral bone formation. We have previously shown that Mmp-13 gene expression is highly induced by PTH treatment in osteoblastic UMR 106-01 cells, as well as primary osteoblasts. Here, we show that p300/CBP-associated factor (PCAF), in addition to p300 and Runx2, is required for PTH activation of Mmp-13 transcription. PCAF was increasingly recruited to the MMP-13 proximal promoter region after PTH treatment, and this was associated with an increase in RNA polymerase II recruitment and histone acetylation. In addition, PTH treatment increased the acetylation of PCAF, a process that required p300. Knockdown of PCAF, p300, or Runx2 by siRNA decreased Mmp-13 mRNA expression after PTH treatment in both UMR 106-01 cells and primary osteoblasts. We found that there is a mutual dependence between p300 and PCAF to be recruited to the Mmp-13 promoter after PTH treatment. In promoter-reporter assays, p300 and PCAF had an additive effect on PTH stimulation of MMP-13 promoter activity, and this required their histone acetyltransferase activity. Our findings demonstrate that PCAF acts downstream of PTH signaling as a transcriptional coactivator that is required for PTH stimulation of MMP-13 transcription. PCAF cooperates with p300 and Runx2 to mediate PTH activation of MMP-13 transcription.

The mechanism by which enzymes recognize the "uniform" collagen triple helix is not well understood. Matrix metalloproteinases (MMPs) cleave collagen after the Gly residue of the triplet sequence Gly∼[Ile/Leu]-[Ala/Leu] at a single, unique, position along the peptide chain. Sequence analysis of types I-III collagen has revealed a 5-triplet sequence pattern in which the natural cleavage triplets are always flanked by a specific distribution of imino acids. NMR and MMP kinetic studies of a series of homotrimer peptides that model type III collagen have been performed to correlate conformation and dynamics at, and near, the cleavage site to collagenolytic activity. A peptide that models the natural cleavage site is significantly more active than a peptide that models a potential but non-cleavable site just 2-triplets away and NMR studies show clearly that the Ile in the leading chain of the cleavage peptide is more exposed to solvent and less locally stable than the Ile in the middle and lagging chains. We propose that the unique local instability of Ile at the cleavage site in part arises from the placement of the conserved Pro at the P(3) subsite. NMR studies of peptides with Pro substitutions indicate that the local dynamics of the three chains are directly modulated by their proximity to Pro. Correlation of peptide activity to NMR data shows that a single locally unstable chain at the cleavage site, rather than two or three labile chains, is more favorable for cleavage by MMP-1 and may be the determining factor for collagen recognition.

The behavioral and psychological symptoms of dementia (BPSD), such as psychosis, agitation, or aggression have a considerable negative impact on the quality of life of both patients and their caregivers. Multiple studies have demonstrated that atypical antipsychotics are efficacious in the treatment of the aggressive and psychotic symptom clusters, and here we review their use in this indication. Because of the safety concerns associated with the use of atypical antipsychotics in this population, these drugs must be used judiciously. For patients with severe BPSD such as psychosis, agitation, or aggression, for whom there are few options, atypical antipsychotics, particularly risperidone and olanzapine, should be considered.

We report observations of three gravitationally lensed supernovae (SNe) in the Cluster Lensing And Supernova survey with Hubble (CLASH) Multi-Cycle Treasury program. These objects, SN CLO12Car (z = 1.28), SN CLN12Did (z = 0.85), and SN CLA11Tib (z = 1.14), are located behind three different clusters, MACSJ1720.2+3536 (z = 0.391), RXJ1532.9+3021 (z = 0.345), and A383 (z = 0.187), respectively. Each SN was detected in Hubble Space Telescope optical and infrared images. Based on photometric classification, we find that SNe CLO12Car and CLN12Did are likely to be Type Ia supernovae (SNe Ia), while the classification of SN CLA11Tib is inconclusive. Using multi-color light-curve fits to determine a standardized SN Ia luminosity distance, we infer that SN CLO12Car was approx. 1.0 +/- 0.2 mag brighter than field SNe Ia at a similar redshift and ascribe this to gravitational lens magnification. Similarly, SN CLN12Did is approx. 0.2 +/- 0.2 mag brighter than field SNe Ia. We derive independent estimates of the predicted magnification from CLASH strong+weak-lensing maps of the clusters (in magnitude units, 2.5 log10 ): 0.83 +/- 0.16 mag for SN CLO12Car, 0.28 +/- 0.08 mag for SN CLN12Did, and 0.43 +/- 0.11 mag for SN CLA11Tib. The two SNe Ia provide a new test of the cluster lens model predictions: we find that the magnifications based on the SN Ia brightness and those predicted by the lens maps are consistent. Our results herald the promise of future observations of samples of cluster-lensed SNe Ia (from the ground or space) to help illuminate the dark-matter distribution in clusters of galaxies, through the direct determination of absolute magnifications.

Most newly synthesized lysosomal enzymes contain a transient carbohydrate modification, mannose 6-phosphate (Man-6-P), which signals their vesicular transport from the Golgi to the lysosome via Man-6-P receptors (MPRs). We have examined Man-6-P glycoproteins in human urine by using a purified soluble fragment of the soluble cation-independent MPR (sCI-MPR) as a preparative and analytical affinity reagent. In a survey of urine samples from seven healthy subjects, the pattern of Man-6-P glycoproteins detected with iodinated sCI-MPR as a probe in a blotting assay was essentially identical in each, regardless of sex or age. Two bands of approx. 100 and 110 kDa were particularly prominent. Man-6-P glycoproteins in human urine were purified by affinity chromatography on immobilized sCI-MPR. Seven distinct bands revealed by SDS/PAGE and Coomassie Blue staining were subjected to N-terminal sequence analysis. The prominent 100 and 110 kDa Man-6-P glycoproteins were identified as N-acetylglucosamine-6-sulphatase and alpha-glucosidase respectively. This identification was confirmed by molecular mass determinations on the two major bands after deglycosylation. Sequence analysis revealed arylsulphatase A and several previously unidentified proteins as minor species. Man-6-P glycoproteins were also purified on an analytical scale to determine the proportion of a number of lysosomal enzyme activities represented by the mannose-6-phosphorylated forms. The lysosomal enzymes in urine containing the highest proportion of mannose-6-phosphorylated form were beta-mannosidase (82%), hexosaminidase (27%) and alpha-glucosidase (24%). The profiles of Man-6-P glycoproteins detected by blotting in urine and plasma were not similar, suggesting that the urinary species are not derived from the bloodstream.

BACKGROUND: Glutamate signaling activates MAPK and PI3K/AKT pathways in tumor cells. Treatment with riluzole, a glutamate release inhibitor, has been previously shown to be safe in melanoma patients and produced biologic effects, but did not lead to radiographic responses, possibly due to poor pharmacokinetic properties. Therefore, we conducted a phase Ib trial to determine the safety and tolerability of the combination of the riluzole prodrug troriluzole (BHV-4157, trigriluzole) and the PD-1 antibody nivolumab in patients with advanced solid tumors. METHODS: Patients with advanced or refractory solid tumors and measurable disease per RECIST 1.1 were treated with increasing doses of troriluzole using a semi-Bayesian modified toxicity probability interval dose escalation procedure. Troriluzole monotherapy was orally self-administered for a 14-day lead-in period followed by continuation of troriluzole in combination with nivolumab 240 mg IV every 2 weeks. Endpoints included safety, pharmacokinetics (PK) and efficacy. RESULTS: We enrolled 14 patients with advanced solid tumors (melanoma = 3, NSCLC = 3, renal cell carcinoma = 2, bladder/urothelial = 2, ovarian cancer = 1, adenoid cystic carcinoma = 1, pleural mesothelial = 1, head and neck cancer = 1). Eleven patients had cancer progression on prior therapy with PD-1 or PD-L1 agent. Patients received troriluzole total daily doses from 140 to 560 mg (divided). The most common treatment-related adverse events (TRAE) occurring in ≥ 5 patients (> 35%) were transaminitis and increased lipase. DLT (dose-limiting toxicity) occurred in 3 patients: (1) grade 3 anorexia, (2) grade 3 fatigue and, (3) grade 3 atrial fibrillation. Six patients were treated at the MTD (maximum tolerated dose). No subjects discontinued treatment due to AEs. One response occurred (7%), which was a partial response in a subject who had PD-1 refractory disease. The 6-month PFS rate was 21%. PK data showed that the prodrug troriluzole was efficiently cleaved into riluzole by 2-h post-dosing in all dose cohorts tested. CONCLUSION: The combination of troriluzole and nivolumab was safe and well-tolerated. The MTD of troriluzole was determined to be 420 mg total daily dose. The observed antitumor activity, primarily disease stabilization, is of interest in patients with PD-1 resistant tumors. Trial Registration ClinicalTrials.gov Identifier NCT03229278.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and its primary oxidation product, 1-methyl-4-phenyl-2,3-dihydropyridinium (MPDP+), are mechanism-based inhibitors of monoamine oxidases A and B. The pseudo-first-order rate constants for inactivation were determined for various analogues of MPTP and MPDP+ and the concentrations in all redox states were measured throughout the reaction. Disproportionation was observed for all the dihydropyridiniums, but non-enzymic oxidation was insignificant. The dihydropyridiniums were poor substrates for monoamine oxidase A and, consequently, inactivated the enzyme only slowly, despite partition coefficients lower than those for the tetrahydropyridines. For monoamine oxidase B, the dihydropyridiniums were more effective inactivators than the tetrahydropyridines. Substitutions in the aromatic ring had no major effect on the inactivation of monoamine oxidase B, but the 2'-ethyl- and 3'-chloro-substituted compounds were very poor mechanism-based inactivators of monoamine oxidase A. It is clear that both oxidation steps can generate the reactive species responsible for inactivation.

BACKGROUND: Ethanol intake and preference differences between the selectively bred alcohol-preferring (P) and nonpreferring (NP) rats have generally been studied in a continuous-access paradigm using 10% ethanol. Little is known about the consumption of lower concentrations of ethanol in these lines or consumption of a wide range of ethanol concentrations in limited-access paradigms. Recently, limited-access paradigms have been used to study the biological and pharmacological mechanisms of ethanol consumption in animal models. Such research would be informed by studies investigating ethanol oral self-administration within a limited-access context. Therefore, the current study addressed P, NP, and Sprague-Dawley (SD) rats' consumption of a wide range of ethanol concentrations in a 2-bottle-choice, limited-access procedure. METHODS: Male P, NP, and SD rats were given concurrent access to water and ethanol solutions for 1 h/d, 7 d/wk. Ethanol solutions were presented in an ascending series ranging from 0.01 to 20% (v/v) over 55 days. Ethanol intakes (g/kg), volumes of solutions consumed (mL/kg), and preference ratios were assessed for each rat line at each concentration. RESULTS: Clear differences among the 3 types of rats emerged at an ethanol concentration of 4%, although differences between P and NP rats emerged at concentrations as low as 1.8%. Alcohol-preferring rats almost exclusively preferred ethanol solutions over water at ethanol concentrations of 4% and above, whereas SD and NP rats' preference ratios were more variable. CONCLUSIONS: The results suggest that differences between P and NP rats exist at ethanol concentrations lower than those previously studied in continuous-access paradigms. They also provide a current description of the ranges of ethanol concentrations preferred by P, NP, and SD rats.

The SCAN or leucine-rich domain has been characterized as a highly conserved sequence in zinc finger transcription factors that mediates selective dimer formation between SCAN-domain-containing proteins. In order to accommodate various SCAN-domain sequence features, a minimal functional folding unit was defined on the premise of proper structural folding and biochemical binding. The 58-amino acid minimal functional units derived from each of four SCAN-domain protein families were subjected to a three-dimensional position-specific scoring matrix (3D-PSSM) and ungapped threading analysis. The resulting fold prediction represented the SCAN-domain's minimal functional unit as a bundle of three alpha helices folded to a core structure. In addition, the minimal functional folding unit biochemically retained the selective dimerization properties of the native proteins. In order to elucidate the structural components within the SCAN-domain that engage in binding interactions, we attempted to correlate the physicochemical helix properties, as represented by a hydropathy profile, with the experimental dimerization selectivities. The amino-terminal helix revealed the highest diversity measure among the three helices of the minimal functional unit and is therefore likely to offer critical surface-exposed binding residues. Indeed, by interchanging the amino-terminal helix between SCAN-domains without alteration of their structural frames consisting of conserved hydrophobic residues, a modulation of binding preferences was demonstrated. The minimal functional folding unit of SCAN-domains may therefore contain within the amino-terminal alpha helix structural components that determine selective dimerization patterns and combinatorial control of transcription factors.

Abstract Caregivers can make a difference in the quality of life of residents in long-term care facilities (LTC) through the application of behavioral management techniques. This is widely accepted, but has little supporting data. In reality, a clear definition of the practice of caregiving in these facilities does not yet exist. Here, we present an overview of extant training in an LTC facility, identify problems, and point to core modules for such training. We also discuss quality of care in these settings. We discuss studies that support these ideas. We then discuss what we call the necessary ingredients of care. These involve the caregiver, the resident, and the system. Importantly, all must be committed for quality indicators to change. Key Words: Nursing homeslong-term care facilitiestrainingelderlycaregiving

Telemarketing is a growth industry that falls within the realm of information management. The telemarketing services provided by AT&T American Transtech grew from non-existence to a position of leadership in the industry in just four years. The organization was able to accomplish this feat with a total systems approach using innovative methods for approaching and satisfying new markets. Hardware innovations, including the use of ISDN, along with a humanized implementation have been combined in this successful effort. This article describes the telemarketing service effort of American Transtech and the lessons learned that could be applied to other information management organizations.

Nausea and vomiting are common complaints in the postoperative period and contribute to patient distress and delay of discharge for outpatient surgical procedures. Laparoscopic procedures are associated with a high incidence of postoperative nausea and vomiting (PONV) episodes. Parenteral use of metoclopramide prevents and treats PONV. The intranasal route provides rapid and complete absorption of metoclopramide without many of the adverse effects observed with parenteral administration of the drug. We performed a prospective, double-blinded, randomized, placebo-controlled study to evaluate the safety and efficacy of metoclopramide 20 mg administered intranasally for emetic prophylaxis in laparoscopic surgery patients. The results from 109 patients enrolled in the study showed that this intranasal dose of metoclopramide may be ineffective in preventing the occurrence of PONV. The poor performance of the intranasal metoclopramide formulation in this study cannot be attributed to patient-specific and perioperative factors. It may be due to an inadequate dose or slow absorption of the drug. The small sample size, however, may also have been a factor.

Abstract: Previous work has shown that incubation of hippocampal slices in medium without added calcium markedly attenuates the capacity of vasoactive intestinal peptide (VIP) to elevate cyclic AMP levels. The present studies examined the mechanism that confers calcium dependence on VIP stimulation of cyclic AMP accumulation in hippocampal slices. Calcium dependence was apparent immediately on slice preparation and was reversible only if calcium ions were added back very early during slice incubation (within 5 min). The cyclic AMP response to VIP was not abolished by preincubating slices in 100 μ M adenosine, suggesting that calcium‐dependent, VIP‐induced release of adenosine does not mediate VIP elevation of cyclic AMP. VIP‐stimulated cyclic AMP accumulation was not decreased by agents that block calcium influx (verapamil, nifedipine, magnesium ions), or by calmodulin antagonists (trifluoperazine, calmidozolium). In fact both verapamil (100 μ M ) and magnesium (14 m M ) augmented VIP stimulation of cyclic AMP generation. Incubation of slices with the phosphodiesterase inhibitor 1‐methyl‐3‐isobutylxanthine (MIX) did not affect VIP activation of cyclic AMP accumulation if slices were incubated without added calcium, but MIX did enhance VIP elevation of cyclic AMP content in slices incubated with calcium. Thus calcium dependence of the cyclic AMP response to VIP in hippocampal slices is unlikely to result from VIP‐dependent calcium influx, from interactions with calmodulin, or from calcium‐inhibited phosphodiester‐ase(s). The accumulation of cyclic AMP induced by two other agonists, norepinephrine (NE) and forskolin, was also dependent on the presence of calcium ions in the bathing medium. The data are also consistent with the hypothesis that VIP, NE, and forskolin stimulate cyclic AMP accumulation in hippocampal slices by interacting with the calcium‐dependent adenylate cyclase.