Fat Mass Is Negatively Associated with Muscle Strength and Jump Test Performance.

BACKGROUND: It is known that maintenance of muscle mass cannot prevent loss of muscle strength in older adults. Recent evidence suggests that fat mass can weaken the relationship between muscle mass and functional performance. No information exists if fat mass can independently affect muscle strength and jump test performance in middle-aged and older adults. OBJECTIVE: To assess the independent relationships between fat mass, leg muscle mass, lower extremity muscle strength, and jump test performance in adults, 55-75 years of age. DESIGN: Cross-sectional. SETTING: University laboratory. PARTICIPANTS: Fifty-nine older adults (men, n = 27, age = 64.8 ± 6.5 years; women, n = 32, age = 62.5 ± 5.1 years) participated in this study. MEASUREMENTS: Dual energy X-ray absorptiometry was used to measure fat mass and leg muscle mass. An average of 3 maximal countermovement jumps was used to calculate jump power and jump height. Two leg press and hip abduction strength were assessed by 1-repetition maximum testing. RESULTS: Stepwise sequential regression analysis of fat mass and leg muscle mass versus jump test performance and measures of muscle strength after adjusting for age, height, and physical activity revealed that fat mass was negatively associated with jump height (p = 0.047, rpartial = -0.410) in men. In women, fat mass was negatively associated with jump height (p = 0.003, rpartial = -0.538), leg press (p = 0.002, rpartial = -0.544), and hip abduction strength (p < 0.001, rpartial = -0.661). Leg muscle mass was positively associated with jump power in women (p = 0.047, rpartial = 0.372) only. CONCLUSIONS: Fat mass has an independent negative relationship with jump test performance in middle-aged and older men and women. This has clinical implications for rehabilitating neuromuscular performance in middle-aged and older adults.

Antimicrobial susceptibility and minimal inhibitory concentration of Pseudomonas aeruginosa isolated from septic ocular surface disease in different animal species.

The purpose of this study was to evaluate the antibiotic susceptibility profile of Pseudomonas aeruginosa isolated from different animal species with septic ocular surface disease. Sixteen strains of P. aeruginosa were isolated from different species of animals (dog, cat, horse, penguin and brown bear) with ocular surface diseases such as conjunctivitis, keratocojnuctivits sicca and ulcerative keratitis. These isolates were tested against 11 different antimicrobials agents using the Kirby-Bauer disk-diffusion method. Minimum inhibitory concentrations (MICs) were determined using E-tests for two antibiotics (tobramycin and ciprofloxacin) commonly used in veterinary ophthalmology practice. Imipenem was the most effective antibiotic, with 100% of the strains being susceptible, followed by amikacin (87.5%), gentamicin, norfloxacin, gatifloxacin and polymyxin (both with 81.5%of susceptibility). MIC90 of ciprofloxacin was 2 µg/ml and the values found ranged from 0.094 µg/ml to 32 µg/ml. For tobramycin, MIC90 was 32 µg/ml and ranged from 0.25 µg/ml to 256 µg/ml. The most effective in vitro antibiotic tested against P. aeruginosa in this study was imipenem, followed by amikacin. The 3 mg/ml eye drops commercially available ciprofloxacin presentations were in vitro effective against all strains tested in this study if applied up to 4 hours after instillation. Whereas for tobramycin the 3 mg/ml eye drops commercial presentations were not in vitro effective against some strains isolated in this study. Thus for ocular infections with P. aeruginosa when using tobramycin the ideal recommendation would be to either use eye drops with higher concentrations or decrease the frequency intervals from four to a minimum of every two hours.

Modulation of gastrointestinal function by MuDelta, a mixed µ opioid receptor agonist/ µ opioid receptor antagonist.

BACKGROUND & PURPOSE: Loperamide is a selective µ opioid receptor agonist acting locally in the gastrointestinal (GI) tract as an effective anti-diarrhoeal but can cause constipation. We tested whether modulating µ opioid receptor agonism with δ opioid receptor antagonism, by combining reference compounds or using a novel compound ('MuDelta'), could normalize GI motility without constipation. EXPERIMENTAL APPROACH: MuDelta was characterized in vitro as a potent µ opioid receptor agonist and high-affinity δ opioid receptor antagonist. Reference compounds, MuDelta and loperamide were assessed in the following ex vivo and in vivo experiments: guinea pig intestinal smooth muscle contractility, mouse intestinal epithelial ion transport and upper GI tract transit, entire GI transit or faecal output in novel environment stressed mice, or four weeks after intracolonic mustard oil (post-inflammatory). Colonic δ opioid receptor immunoreactivity was quantified. KEY RESULTS: δ Opioid receptor antagonism opposed µ opioid receptor agonist inhibition of intestinal contractility and motility. MuDelta reduced intestinal contractility and inhibited neurogenically-mediated secretion. Very low plasma levels of MuDelta were detected after oral administration. Stress up-regulated δ opioid receptor expression in colonic epithelial cells. In stressed mice, MuDelta normalized GI transit and faecal output to control levels over a wide dose range, whereas loperamide had a narrow dose range. MuDelta and loperamide reduced upper GI transit in the post-inflammatory model. CONCLUSIONS AND IMPLICATIONS: MuDelta normalizes, but does not prevent, perturbed GI transit over a wide dose-range in mice. These data support the subsequent assessment of MuDelta in a clinical phase II trial in patients with diarrhoea-predominant irritable bowel syndrome.

Anti-inflammatory role of glycine in reducing rodent postoperative inflammatory ileus.

BACKGROUND: Inflammatory events within the intestinal muscularis, including macrophage activation and leukocyte recruitment, have been demonstrated to participate in causing postoperative ileus. Recently, glycine has gained attention due to its beneficial immunomodulatory effects in transplantation, shock and sepsis. METHODS: Muscularis glycine receptors were investigated by immunohistochemistry. Gastrointestinal motility was assessed by in vivo transit distribution histograms with calculated geometric center analysis and jejunal circular smooth muscle contractility in a standard organ bath. The impact of glycine on the muscularis inflammatory responses to surgical manipulation of the intestine were measured by real-time PCR, nitric oxide Griess reaction, prostaglandin ELISA, Luminex and histochemistry. KEY RESULTS: Glycine-gated chloride channels were immunohistochemically localized to muscularis macrophages and postoperative infiltrating leukocytes. Preoperative glycine treatment significantly improved postoperative gastrointestinal transit and jejunal circular muscle contractility. Preoperative glycine injection significantly reduced the induction of interleukin-6 (IL-6), tumor necrosis factor-α, inducible nitric oxide synthase and intercellular adhesion molecule-1 mRNAs, which was associated with the attenuation in postoperative leukocyte recruitment. Nitric oxide and prostanoid release from the postsurgical inflamed muscularis was diminished by glycine. The secretion of the inflammatory proteins IL-6, monocyte chemotactic protein-1/chemokine ligand 2 and macrophage inflammatory protein-1α/chemokine ligand 3 were also significantly decreased by glycine pretreatment. CONCLUSIONS & INFERENCES: The data indicate that preoperative glycine reduces postoperative ileus via the early attenuation of primal inflammatory events within the surgically manipulated gut wall. Therapeutic modulation of resident macrophages by glycine is a potential novel pharmacological target for the prevention of postoperative ileus.

Response to hepatitis A vaccine in HIV-positive patients.

The USPHS/IDSA guidelines for Prevention of Opportunistic Infections in Persons with human immunodeficiency virus (HIV) recommends that all susceptible HIV+ patients at increased risk for hepatitis A virus (HAV) or with chronic liver disease, be vaccinated against HAV. Immune response to HAV vaccine has not been well studied in HIV+ patients. In particular, there is little information in the literature regarding the effect and relationship of the CD4 count and the immune response in HIV patients. A retrospective analysis of HIV+ patients who received HAV vaccine was performed, and the antibody response to HAV (anti-HAV) measured. Univariate and multivariate analyses were performed to determine predictors of response to vaccine administration. Of the 503 patients evaluated, 138 patients completed their HAV vaccination series and 48% of them had postvaccine anti-HAV positive results (responders). There was no difference in age, race, antiretroviral therapy use, or hepatitis C virus exposure between responders and nonresponders. In univariate analysis, responders were more likely to be female (40.3%vs 21.1%, P = 0.01), have a higher CD4 count at vaccine (508.6 cells/mm3 vs 344.3 cells/mm3, P = 0.001) and marginally lower viral load at vaccine (2.65 log copies vs 2.94 log copies, P = 0.07). Multivariate analysis showed that female gender and higher CD4 count at vaccine were independent predictors of response to vaccine. Forty-eight per cent of our HIV+ patients responded to HAV vaccine administration. This is much lower than reported rates of 100% in HIV-negative patients. Female gender and CD4 count at vaccine, but not CD4 nadir, predicted response to vaccine.

Immunomodulatory effects of inhaled carbon monoxide on rat syngeneic small bowel graft motility.

BACKGROUND: Intestinal transplantation provokes an intense inflammatory response within the graft muscularis that causes intestinal ileus. We hypothesised that endogenously produced anti-inflammatory substances could be utilised as novel therapeutics. Therefore, we tested the protective effects of inhaled carbon monoxide (CO) and an endogenous haeme oxygenase 1 (HO-1) anti-inflammatory mediator on transplant induced inflammatory responses and intestinal ileus in the rat. METHODS: Gastrointestinal transit of non-absorbable FITC labelled dextran and in vitro jejunal circular muscle contractions were measured in controls and syngeneic orthotopic transplanted animals with and without CO inhalation (250 ppm for 25 hours). Inflammatory mRNAs for interleukin (IL)-6, IL-1beta, tumour necrosis factor alpha (TNF-alpha), intercellular adhesion molecule 1 (ICAM-1), inducible nitric oxide (iNOS), cyclooxygenase 2 (COX-2), and IL-10 were quantified by real time reverse transcriptase-polymerase chain reaction and HO-1 by northern blot. Histochemical stains characterised neutrophil infiltration and enterocyte apoptosis. RESULTS: Transplantation delayed transit and suppressed jejunal circular muscle contractility. Transplantation induced dysmotility was significantly improved by CO inhalation. Transplantation initiated a significant upregulation in IL-6, IL-1beta, TNF-alpha, ICAM-1, iNOS, COX-2, and HO-1 mRNAs with the graft muscularis. CO inhalation significantly decreased expression of IL-6, IL-1beta, iNOS, and COX-2 mRNAs. CO also significantly decreased serum nitrite levels (iNOS activity). CONCLUSIONS: CO inhalation significantly improved post-transplant motility and attenuated the inflammatory cytokine milieu in the syngeneic rat transplant model. Thus clinically providing CO, the end product of the anti-inflammatory HO-1 pathway, may prove to be an effective therapeutic adjunct for clinical small bowel transplantation.

Marijuana abstinence effects in marijuana smokers maintained in their home environment.

BACKGROUND: Although withdrawal symptoms are commonly reported by persons seeking treatment for marijuana dependence, the validity and clinical significance of a marijuana withdrawal syndrome has not been established. This controlled outpatient study examined the reliability and specificity of the abstinence effects that occur when daily marijuana users abruptly stop smoking marijuana. METHODS: Twelve daily marijuana smokers were assessed on 16 consecutive days during which they smoked marijuana as usual (days 1-5), abstained from smoking marijuana (days 6-8), returned to smoking marijuana (days 9-13), and again abstained from smoking marijuana (days 14-16). RESULTS: An overall measure of withdrawal discomfort increased significantly during the abstinence phases and returned to baseline when marijuana smoking resumed. Craving for marijuana, decreased appetite, sleep difficulty, and weight loss reliably changed across the smoking and abstinence phases. Aggression, anger, irritability, restlessness, and strange dreams increased significantly during one abstinence phase, but not the other. Collateral observers confirmed participant reports of these symptoms. CONCLUSIONS: This study validated several specific effects of marijuana abstinence in heavy marijuana users, and showed they were reliable and clinically significant. These withdrawal effects appear similar in type and magnitude to those observed in studies of nicotine withdrawal.

Do drivers drink more when they use a safe ride?

PURPOSE: Offering drinkers free safe rides (SRs) home can prevent DWI, but some suggest that it may also promote excessive drinking. METHODS: Forty-two respondents to surveys completed by 472 drinkers in barrooms reported that they used a SR during the first 9 months they were offered. RESULTS: Their usual estimated blood alcohol concentration (BAC) was significantly greater when they used a SR than on all occasions when they drank outside their homes. However, only 24% of respondents increased their usual BAC from a " low risk" category (BAC <0.10) when drinking outside the home, to a "high risk" category (BAC > or = 0.10) when using a SR. The majority (72%) of respondents did not change their BAC categories. One decreased his/her BAC from > or = 0.10 when drinking outside the home to <0.10 when using a SR. IMPLICATIONS: These findings, coupled with data showing that respondents tended to use SRs infrequently (3.69 times per year), reduce concerns about the mean increase in their BACs when using SRs. Findings do not, however, resolve the issue of whether respondents whose drinking increased when they used a SR did so because a SR was available, or decided not to drive because they drank more. Recommendations for improving SR programs are discussed.

Carbon dioxide laser skin resurfacing with a cooled handpiece.

BACKGROUND AND OBJECTIVE: Carbon dioxide (CO(2)) laser skin resurfacing has become an accepted procedure for the rejuvenation of aged and wrinkled facial skin. The benefits of this technique have been limited by the prolonged postoperative erythema, delayed re-epithelialization, and potential changes in pigmentation. We investigated the effects of coupling a pulsed CO(2) laser to a zinc selenide handpiece at various temperatures to cool and conduct heat from the skin surface. We compared the heat conducting handpiece to standard techniques used to determine the effects on epithelial preservation, depth of thermal damage, and new collagen growth. STUDY DESIGN/MATERIALS AND METHODS: The dorsal aspect of piglets was treated with four handpiece arrangements: no thermally conducting window; a zinc selenide thermally conducting window at room temperature; and the zinc selenide thermally conducting window cooled to 10 and 5 degrees C. Intensities were measured to account for reflections from the surface of the handpiece. With the CO(2) laser in scanning mode and a spot size of 1 mm, each handpiece was used at intensities ranging from 8-30 W in one pass. Biopsies for histopathologic analysis and determination of the degree of epithelial damage and the depth of thermal damage were taken on days 0, 2, and 16. RESULTS: All specimens revealed complete re-epithelialization by day 16. The addition of a cooled handpiece did not affect either the amount of epithelial preservation or the depth of thermal damage. CONCLUSIONS: There is no significant advantage in using a cooled or room temperature zinc selenide handpiece for epithelial preservation or decreasing the inflammatory response in CO(2) laser resurfacing.

Rubber band ligation for hemorrhoidal disease can be safely performed in select HIV-positive patients.

PURPOSE: Many surgeons have abandoned the use of rubber band ligation for the treatment of hemorrhoids in patients infected with human immunodeficiency virus because of the belief that this procedure could lead to disastrous outcomes. This study was designed to evaluate the safety and efficacy of rubber band ligation in otherwise healthy human immunodeficiency virus-positive patients. METHODS: A retrospective chart review of healthy human immunodeficiency virus-positive patients who underwent rubber band ligation for symptomatic hemorrhoids between April 1993 and May 2000 was conducted. RESULTS: The study group comprised 11 patients. All patients were male, with a median age of 48 (range, 32-64) years. Mean T-cell helper count was 450 (range, 200-1,000) cells/microl. A median of 2 (range, 1-4) rubber band ligations were performed per patient. The median length of follow-up was seven (range, 1-28) months. There were no deaths or complications in any study group patient. Eight patients (73 percent) had excellent results, with complete resolution of symptoms. Two patients (18 percent) had initial improvement but subsequently had hemorrhoidectomy because of recurrent symptoms. Only one patient (9 percent) had no benefit from rubber band ligation and underwent hemorrhoidectomy. CONCLUSION: These data suggest that asymptomatic human immunodeficiency virus-positive patients can be treated safely and effectively with rubber band ligation for symptomatic hemorrhoids.

Membrane type I-matrix metalloproteinase-mediated degradation of type I collagen by oral squamous cell carcinoma cells.

Oral squamous cell carcinomas are highly invasive lesions that destroy adjacent tissues and invade bone and muscle, which is most likely the result of matrix metalloproteinase (MMP) activity. We examined three cell lines derived from squamous cell carcinoma of the tongue for their intrinsic capacities to degrade interstitial collagen with the goal of identifying the matrix-degrading enzymes. SCC-25 and SCC-15 cells degrade reconstituted fibrillar type I collagen in the absence of exogenous growth factors or cytokines when seeded as a colony on dried films. Degradation is confined to the subjacent matrix, is enhanced 2-3-fold by phorbol ester, and is strictly MMP-dependent, as it is blocked by BB-94 and tissue inhibitor of metalloproteinases-2 but not by inhibitors of serine and cysteine proteinases. Both cell lines express active (M(r) 57,000) membrane type I-MMP (MT1-MMP) on their surfaces, as detected by surface biotinylation and immunoprecipitation. Concomitantly, both cell lines activate endogenous MMP-2 when cultured on type I collagen films, as assessed by zymography. Phorbol ester treatment enhances collagen-induced MMP-2 activation, which is accompanied by the appearance of a surface-labeled M(r) 43,000 form of MT1-MMP. Treatment of cells with a synthetic furin inhibitor, which inhibits processing of the MT1-MMP zymogen, blocks collagen degradation. In contrast, CAL 27 cells do not degrade collagen under either basal or phorbol 12-myristate 13-acetate-stimulated conditions. Although proMT1-MMP (M(r) 63,000/65,000) is detectable in these cells by immunoblot analysis, they express greatly reduced levels of active MT1-MMP on their surfaces relative to SCC-25 and SCC-15 cells. Correspondingly, CAL 27 cells cultured on collagen express neither latent nor active gelatinases. Immunoblots of lysates and conditioned media revealed the constitutive expression of proMMP-1 and proMMP-13 in all three cell lines. We conclude that in the absence of exogenous growth factors or accessory stromal cells, degradation of interstitial collagen by oral squamous cell carcinoma cells requires a threshold level of active MT1-MMP on cell surfaces.

Conserved function of Caenorhabditis elegans UNC-30 and mouse Pitx2 in controlling GABAergic neuron differentiation.

We are taking a cross-species approach to identify genes that are required for mammalian GABAergic neuron differentiation. On the basis of homeodomain similarity, the vertebrate Pitx genes appear to be orthologs of unc-30, a Caenorhabditis elegans gene necessary for differentiation of the GABAergic phenotype of type D neurons. One of the Pitx genes, Pitx2, is expressed in regions of GABAergic neurogenesis in the mammalian brain. These observations led us to test the functional conservation of the mouse Pitx2 and worm unc-30 genes using a rescue assay. Pitx2 rescues the GABAergic differentiation defect and partially rescues the axon guidance and behavioral phenotypes of unc-30 mutants, indicating a high degree of functional conservation between these evolutionarily related genes. Previous studies show that UNC-30 directly regulates the unc-25/glutamate decarboxylase gene that encodes the enzyme for GABA synthesis. We find that the promoter regions of the mouse and human genes coding for the 67 kDa glutamate decarboxylase (Gad1) also contain binding sites matching the UNC-30/Pitx2 consensus binding site sequence. We show that these sites specifically bind to Pitx2 protein in vitro and that in transfected neuroblastoma cells, the Pitx2 binding sites contribute to the basal activity of the Gad1 promoter. Furthermore, in cotransfection experiments, we find that Pitx2 strongly activates the Gad1 promoter. These results indicate that Pitx2 may regulate Gad1 expression in mammals, suggesting a new role for this key developmental transcription factor as a regulator of GABAergic differentiation during mammalian neural development. Our results suggest that some of the mechanisms regulating GABAergic differentiation are evolutionarily conserved.

DWI prevention: profiles of drinkers who use designated drivers.

The effectiveness of designated driver (DD) use in preventing driving while intoxicated (DWI) depends on whether drinkers at risk for DWI use DDs. Bivariate and logistic regression analyses conducted on data from 1,391 Computer-Assisted Telephone Interviews (CATIs) and from 902 barroom patron surveys showed that DD users, compared to nonusers, tended to be at-risk, heavier drinkers. For example, logistic regression using the CATI sample indicated that DD users were more likely to drink more often outside the home, to achieve higher blood alcohol concentrations (BACs) when drinking outside the home, to ride with intoxicated drivers (RID), and to be heavy drinkers on D. Cahalan et al.'s (1969) Quantity-Frequency-Variability (QFV) index. Similarly, logistic regression using the barroom sample showed that DD users tended to be heavy drinkers on the QFV index, and were more likely to drive after drinking and to ride with intoxicated drivers. Additional analyses showed that DD users also were more likely than nonusers to engage in other behavior to avoid DWI, such as drinking less, waiting to drive until the effects of alcohol diminish, walking home, and staying overnight. These results are consistent with other findings from a related study by the current authors which showed that at-risk drinkers also used free safe (taxi) rides to avoid DWI, however were still more likely to report DWI and RID behavior (B. D. Caudill, W. M. Harding, & B. Moore, in press). Consequently, DWI prevention efforts may be improved by future research aimed at learning why such at-risk drinkers sometimes take steps to avoid DWI and sometimes do not.

Receptor mediated uptake of peptides that bind the human transferrin receptor.

A biopanning process designed to find peptide epitopes specific for cell surface receptors has been used in this study to select seven- and 12-amino-acid peptides capable of binding to and internalizing with the human transferrin receptor (hTfR). Through sequential rounds of negative and positive selection, two peptide sequences were identified that specifically bind to the hTfR. Phage containing the sequences HAIYPRH or THRPPMWSPVWP were inhibited from binding the hTfR in a dose-dependent fashion when peptides of the same sequence were present in a competition assay. Interestingly, transferrin did not compete with either of these sequences for receptor binding, suggesting that these peptides bind a site on the hTfR distinct from the transferrin binding site. When either of these sequences was expressed as a fusion to green fluorescent protein (GFP), the recombinant GFP molecule was internalized in cells expressing the hTfR. These studies suggest that the two peptides can be used to target other proteins into the endosomal pathway. Further, they provide a strategy for identifying peptides that bind to other cell surface receptors that can be used for both diagnostic and therapeutic purposes.

Do companions of designated drivers drink excessively?

PURPOSE: A common criticism of designated driver programs (DDPs) is that they promote excessive drinking among companions of the designated driver (DD). METHODS: Data were collected from two representative samples of drinkers using computer-assisted telephone interviews (CATIs), and questionnaires administered to customers in barrooms. RESULTS: Most respondents drank moderately--had usual estimated blood alcohol concentrations (BACs) of less than 0.10 when they used DDs. Differences between respondents' estimated BACs when they used a DD and when they drank outside their homes were very small: 0.017 in both samples. Additional analyses examined shifts between lower and higher categories of risk defined as a BAC of less than 0.10 and a BAC of 0.10 or greater. A minority, 15% of CATI and 30% of barroom respondents, switched to the higher risk category when using a DD. These CATI and barroom respondents increased their BACs by an average of 0.089 and 0.11, respectively. Risk associated with this increase was mitigated, however, by respondents' infrequent use of DDs. IMPLICATIONS: Use of DDs was not generally associated with excessive alcohol consumption. Since a minority of respondents did drink heavily when using a DD, programs promoting DD use should caution drinkers that the availability of a DD is not an excuse for excessive consumption, and remind hosts and servers that they should not overserve their guests or customers even when they have a DD.

Tobacco smoking in marijuana-dependent outpatients.

PURPOSE: Among marijuana-dependent individuals, approximately 50% smoke tobacco. These individuals are exposed to increased risks of respiratory and other health problems. The current study examined whether tobacco smoking among marijuana-dependent individuals is also associated with increased psychosocial and substance abuse problems. METHODS: Marijuana-dependent individuals (N=174) seeking treatment for marijuana problems completed a 2-3 h assessment. Current tobacco smokers were compared to ex-smokers and never smokers on demographic, psychosocial, and substance use characteristics, and treatment outcome. In addition to univariate comparisons, multivariate analyses using multinomial logistic regression were conducted to control for the correlated nature of the predictor variables. RESULTS: Current tobacco smokers earned less income and reported histories of more alcohol problems than never smokers and had fewer years of education, more legal problems, more psychiatric symptoms, and an earlier age of marijuana initiation than ex- and never smokers. Over the course of treatment, current tobacco smokers had significantly fewer marijuana-negative urine specimens and fewer weeks of continuous marijuana abstinence than ex-smokers. IMPLICATIONS: Current tobacco smokers appear to represent a subgroup of marijuana-dependent individuals who have increased psychosocial problems compared to ex- and never smokers and may not respond as well to treatment than ex-smokers.

Neural pathways regulating Brunner's gland secretion in guinea pig duodenum in vitro.

This study examined the neural pathways innervating Brunner's glands using a novel in vitro model of acinar secretion from Brunner's glands in submucosal preparations from the guinea pig duodenum. Neural pathways were activated by focal electrical stimulation and excitatory agonists, and videomicroscopy was used to monitor dilation of acinar lumen. Electrical stimulation of perivascular nerves evoked large dilations that were blocked by TTX (1 microM) or the muscarinic receptor antagonist 4-diphenylacetoxy-N-(2-chloroethyl)-piperidine hydrochloride (1 microM). The nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium iodide (100 microM) had no effect, and the nerve-evoked responses were not inhibited by hexamethonium (200 microM). Dilations were abolished in preparations from chronically vagotomized animals. Activation of submucosal ganglia significantly dilated submucosal arterioles but not Brunner's glands. Effects of electrical stimulation of perivascular and submucosal nerves were not altered by guanethidine. Capsaicin and substance P also dilated arterioles but had no effect on Brunner's glands. Cholinergic (choline acetyltransferase-immunoreactive) nerve fibers were found in Brunner's glands. These findings demonstrate that Brunner's glands are innervated by cholinergic vagal fibers but not by capsaicin-sensitive or intrinsic enteric nerves.

Induction of collagenase-3 (MMP-13) in rheumatoid arthritis synovial fibroblasts.

There is a growing body of evidence that implicates matrix metalloproteinases (MMPs) as major players in numerous diseased conditions. The articular cartilage degradation that is characteristic of rheumatoid arthritis (RA) is believed to be mediated by the collagenase subfamily of matrix metalloproteinases. The preference of collagenase-3 (CL-3) for collagen type II makes it a likely candidate in the turnover of articular cartilage and a potential target for drug development. In this study, RA synovial membrane tissue was shown to express CL-3 mRNA by reverse transcriptase-polymerase chain reaction (RT-PCR) and protein by immunohistochemistry. Fibroblasts isolated and cultured from RA synovial membrane tissue were induced to express CL-3 mRNA. CL-3 mRNA was detected after PMA treatment in 16 of the 18 RA synovial membrane fibroblast cell lines established for this study. These fibroblasts also expressed mRNA for collagenase-1 (CL-1, MMP-1), membrane type-1 matrix metalloproteinase, gelatinase A, gelatinase B, stromelysin-1, stromelysin-2, TIMP-1, and TIMP-2. They were further shown to express CL-1 mRNA constitutively and CL-3 mRNA only after stimulation with PMA, IL-1, TGF-beta1, TNF-alpha, or IL-6 with IL-6sR. These fibroblasts also expressed after induction both CL-1 and CL-3 at the protein level as determined by Western blot analyses and immunofluorescence.