[Analysis of the effectiveness of coracoid osteotomy and concentric coaxial reconstruction of the glenoid cavity in the treatment of recurrent anterior dislocation of the shoulder joint].

Objective: To investigate the clinical efficacy of the modified Latarjet procedure in the treatment of recurrent anterior subluxation of the shoulder by "coaxial co-arc" reconstruction of the glenoid cavity. Methods: The clinical data of 103 cases (106 shoulders) of recurrent anterior dislocation of the shoulder admitted to the First Affiliated Hospital of the Army Military Medical University from January 2005 to December 2020 were retrospectively studied. Out of these cases, 84 were males and 19 were females; 31 with left-sided injuries while 75 with right-sided injuries, with a mean age of (29.4±11.5) years (16-61 years). The preoperative anterior fear test was positive, and a modified Latarjet procedure was used to reconstruct the shoulder glenoid defect through a "coaxial co-arc". The Rowe score, simple shoulder test (SST) score, and Visual analogue scale (VAS) score of pain were used to assess the shoulder's function. Parameters such as the postoperative shoulder recurrent dislocation rate, shoulder body external rotation angle, and subscapularis muscle strength changes were recorded postoperatively. Moreover, radiographs and CT scans were used to check for the incidence of osteoarthritis (Samson-Prieto score). Results: After a mean follow-up of 9.0 years (1 to 16 years), bony healing occurred 3 to 6 months postoperatively. The Rowe score improved from 40.4±6.5 preoperatively to 93.2±2.5 (P<0.001), the SST score improved from 5.2±1.3 preoperatively to 10.1±1.5 (P<0.001), and the VAS pain score decreased from 3.5±1.9 preoperatively to 1.1±1.2 (P<0.001) at the final follow-up. The angle of lateral external rotation of the shoulder joint was 58.8°±15.6° preoperatively and 57.6°±14.5° postoperatively with no statistically significant difference (P>0.05). There was no statistically significant difference in the measurement of subscapularis muscle strength between the healthy side and the affected side (P>0.05). In 89.6% of patients after surgery, coaxial co-arc reconstruction of the shoulder glenoid was obtained, and the shoulder glenoid defect and postoperative inclusion angle were significantly improved compared with those before surgery (P<0.001). Postoperatively, new-onset osteoarthritis developed in 7 cases (7/98), arthritis progressed in 2 cases (2/8), incisional healing was poor in 2 cases (2/98), and revision surgery was performed in 2 cases (2/98) due to bone mass detachment. Conclusion: Coracoid osteotomy and concentric coaxial reconstruction of the glenoid cavity elicits adequate good clinical efficacy for cases of recurrent anterior shoulder dislocation, with low recurrence rates, low revision rates and low incidence of osteoarthritis.

[Mid-and-long term follow-up of autogenous tibial periosteum-bone complex transplantation for treatment of cystic osteochondral lesion of talus].

Objective: To further clarify the mid-and-long term follow-up results of self-designed tibial periosteum-bone complex transplantation in the treatment of Hepple V osteochondral lesion of the talus(OLTs). Methods: The clinical data of 30 patients with Hepple V OLTs who received treatment in the Sports Medicine Center of the First Affiliated Hospital of Army Military Medical University from October 2011 to January 2019 were analyzed. There were 19 males and 11 females with a mean age of (40±11) years. Patients were treated with autogenous tibial periosteum-bone complex transplantation and were followed up for at least 2 years. The Foot and Ankle Outcome Score (FAOS), American Orthopedic Foot and Ankle Society (AOFAS) ankle-hindfoot score, the visual analog scale score (VAS) of pain, the simplified symptomatology evaluation (SSE) and imaging results before the operation and at the follow-up were recorded and compared. Results: The cohort were followed-up for a mean of 63.9 months (range 24-110 months). Twenty-nine (96.7%) patients were satisfied with the curative effect. The FAOS score was improved from 53.5±6.2 preoperatively to 88.4±6.6 at the final follow-up (P<0.001). The AOFAS ankle-hindfoot scale improved from 61.6±8.2 preoperatively to 90.8±6.8 at the last follow-up (P<0.001). The VAS score decreased from 4.3±0.2 preoperative to 0.7±0.7 at the last follow-up (P<0.001). The SSE score was poor in 14 cases (46.7%), average in 16 cases (53.3%) before the operation; and it was excellent in 23 cases (76.7%), good in 6 cases (20%), average in 1 case (3.3%) at the last follow-up. Imaging examination showed cystic change cure rate was 83.3%, cartilage defects were completely infilled with repair tissue, which didn't show any signs of degeneration. However, repair tissue showed varying degrees of heterogeneous signal compared to the normal articular cartilage. Conclusion: The autograft of tibial periosteum-bone complex transplantation is a safe and feasible method for the treatment of osteochondral lesion of the talus in Hepple V type, with good mid-and-long term clinical effect.

[A comparative study of the calcaneal closing-wedge calcaneal osteotomy versus posterior-superior prominence removal in both sides with Haglund syndrome].

Objective: To compare the clinical outcome of removal of calcaneal posterior-superior prominence and that of calcaneal closing-wedge osteotomy for Haglund syndrome. Methods: From February 2009 to July 2014, 36 patients with Haglund syndrome were included.They were divided into two groups, and each group included 18 patients and underwent removal of calcaneal posterior-superior prominence and calcaneal closing-wedge osteotomy respectively.They were evaluated preoperatively and after 6 , 12 months and 24 months postoperatively by American Orthopedic Foot & Ankle Society (AOFAS) score, VAS score, VISA-A questionnaire and Maryland Foot Score.Fowler-Philip angle and calcaneal posterior slope of the two groups were compared preoperatively and after 6 months.All data were analysis utilizing SPSS 18.0. Results: At six months of follow-up, the weight-bearing lateral X-rays reveals that removal of calcaneal posterior-superior prominence did not change Fowler-Philip angle and calcaneal posterior slope and calcaneal closing-wedge osteotomy decreased Fowler-Philip angle and calcaneal posterior slope significantly[from preoperation (56.5±5.4)°, (120.0±1.3)°to postoperation (48.4±4.6)°, (109.0±5.3)°]. At six months of follow-up, the AOFAS score, VAS score, VISA-A questionnaire and Maryland Foot Score were worse in the wedge calcaneal osteotomy group.At twelve months of follow-up, no significant difference (P>0.05)was found between the two groups in terms of VAS score, and Maryland Foot Score, while the AOFAS score, and VISA-A questionnaire in the wedge calcaneal osteotomy group were better than those of posterior-superior prominence removal group.At twenty-four months of follow-up, the AOFAS score, VAS score, VISA-A questionnaire and Maryland Foot Score were better in the wedge calcaneal osteotomy group (P<0.05). Conclusions: Both the two surgical methods are effective for Haglund syndrome.Calcaneal closing-wedge osteotomy decreased Fowler-Philip angle and calcaneal posterior slope of calcaneus and its clinical outcome appears better than that removal of calcaneal posterior-superior prominence.

A functional magnetic resonance imaging study of the neuronal specificity of an acupoint: acupuncture at Rangu (KI 2) and its sham point.

The neuronal specificity of acupoints has not been entirely supported by the results of previous functional magnetic resource imaging studies. This study tested the specificity of an acupoint using right Rangu (KI 2) and its sham acupoint. The results showed specific cerebral response patterns and thus provided the evidence of the existence of acupoint neuronal specificity.

The safety and effectiveness of TM81, a Chinese herbal medicine, in the treatment of type 2 diabetes: a randomized double-blind placebo-controlled trial.

AIM: TM81 (or Tang-Min-Ling-Wan) is a Chinese medicine. Previous studies suggested that this medicine is effective for treating type 2 diabetes. This controlled trial evaluated the safety and effectiveness of TM81 in the treatment of type 2 diabetic patients. METHODS: This study was a large-scale controlled clinical trial to evaluate the safety and effectiveness of TM81 on type 2 diabetes. After a 2-week run-in period, 480 overweight type 2 early-stage diabetic patients [35-65 years old, HbA1c ≥ 7.0%, fasting plasma glucose (FPG) 7.0-13.9 mM or 2 h plasma glucose (PG) > 11.1 mM, body mass index (BMI) ≥ 24 kg/m(2)] were enrolled. These patients were divided into a TM81 group and placebo group in a 3 : 1 ratio. The subjects received 6 g TM81 or placebo, three times daily for 12 weeks. RESULTS: After treatment, the HbA1c decrease was 1.02% in the TM81 group versus 0.47% in the placebo group. The FPG decreased 0.8 ± 0.1 mM in the TM81 group versus an increase of 0.2 ± 0.2 mM in the placebo group. The PG decreased 2.7 ± 0.3 mM in the TM81 group versus a decrease of 0.9 ± 0.4 mM in the placebo group (all p < 0.05). The TM81 was more effective for patients with higher baseline HbA1c levels. The TM81 group also showed improved ß-cell function and increased homeostatic model assessment (HOMA)-ß. In addition, body weight, BMI and waist circumference of subjects in the TM81 group were reduced, and the symptoms related to diabetes were improved. There were no significant differences in the types and frequency of adverse reactions between the two groups. CONCLUSIONS: The data showed that TM81 is effective in controlling blood glucose level and is safe to use in patients with early-stage type 2 diabetes.

Ginseng berry concentrate prevents colon cancer via cell cycle, apoptosis regulation, and inflammation-linked Th17 cell differentiation.

The Asian ginseng root (Panax ginseng C.A. Meyer) is a very commonly used herbal medicine worldwide. Ginseng fruit, including the berry (or pulp) and seed, is also valuable for several health conditions including immunostimulation and cancer chemoprevention. In this study, the anticancer and anti-proliferative effects of the extracts of ginseng berry and seed were evaluated. The ginsenosides in the ginseng berry concentrate (GBC) and ginseng seed extract (GSE) were analyzed. We then evaluated their anti-colorectal cancer potentials, including antiproliferation, cell cycle arrest, and apoptotic induction. Further investigation consisted of the berry's adaptive immune responses, such as the actions on the differentiation of T helper cells Treg, Th1, and Th17. The major constituents in GBC were ginsenosides Re and Rd, which can be compared to those in the root. The GBC significantly inhibited colon cancer cell growth, and its anti-proliferative effect involved mechanisms including G2/M cell cycle arrest via upregulation of cyclin A and induction of apoptosis via regulation of apoptotic related gene expressions. GBC also downregulated the expressions of pro-inflammatory cytokine genes. For the adaptive immune responses, GBC did not influence Th1 and Treg cell differentiation but significantly inhibited Th17 cell differentiation and thus regulated the balance of Th17/Treg for adaptive immunity. Although no ginsenoside was detected in the GSE, interestingly, it obviously enhanced colon cancer cell proliferation with the underlined details to be determined. Our results suggested that GBC is a promising dietary supplement for cancer chemoprevention and immunomodulation.

Baicalein, an enteric microbial metabolite, suppresses gut inflammation and cancer progression in ApcMin/+ mice.

OBJECTIVE: Chronic inflammation is recognized as a risk factor for colorectal cancer (CRC) development. Baicalin (BI), a major constituent in an anti-inflammatory herb Scutellaria baicalensis, can be biotransformed into baicalein (BE) by the intestinal microbiota. We evaluated the anti-inflammation and anti-CRC effects of the metabolite BE. METHODS: The in vitro biotransformation by human intestinal microbiota from BI into BE has been determined with HPLC. Using a gut-specific ApcMin/+ mouse model, the effects of oral BE on the life span, organ index, and tumor multiplicity were evaluated. The expressions of inflammatory cytokines were determined using ELISA. To verify the in vivo data, the anti-inflammatory and antiproliferative effects of BE were determined with an in vitro cell model. RESULTS: HPLC analysis showed that BI was quickly transformed into BE by the intestinal microbiota. Oral BE (30 mg/kg/day) significantly increased the life span, from 125.2 to 218.4 days (P < 0.01%). BE treatment also decreased intestine index and increased spleen index. Compared with the model group, following BE treatment, tumor numbers were significantly reduced in the small intestine and colon (P < 0.01, P < 0.05, respectively). In the gut tissues, BE treatment significantly reduced inflammatory cytokine levels such as IL-1ß, IL-2, IL-6, IL-10, G-CSF, and GM-CSF. In vitro data supported our in vivo results that the anti-CRC effects of BE were via the inhibition of gut inflammation and induction of cancer cell death. CONCLUSION: Our results suggest that the parent compound BI can be quickly converted into its microbial metabolite BE, which has stronger bioactive effects than BI. Baicalein is an active chemopreventive metabolite for inflammatory associated CRC.

Role of intestinal microbiome in American ginseng-mediated colon cancer prevention in high fat diet-fed AOM/DSS mice [corrected].

OBJECTIVE: Chronic intestinal inflammation is a risk factor for colorectal cancer (CRC) initiation and development. Diets that are rich in Western style fats have been shown to promote CRC. This study was conducted to investigate the role of intestinal microbiome in American ginseng-mediated CRC chemoprevention in a mouse model. The population and diversity of enteric microbiome were evaluated after the ginseng treatment. METHODS: Using an azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced gut inflammation and tumorigenesis mouse model, the effects of oral American ginseng on high fat diet-associated enteric pathology were determined. After establishment of a 16S rRNA illumina library from fecal samples, MiSeq sequencing was carried out to reveal the microbial population. The alpha and beta diversities of microbiome were analyzed. RESULTS: American ginseng significantly attenuated AOM/DSS-induced colon inflammation and tumorigenesis by reducing the colitis score and colon tumor multiplicity. The MiSeq results showed that the majority of sequences fell into three phyla: Firmicutes, Bacteroidetes and Verrucomicrobia. Further, two significant abundance shifts at the family level, Bacteroidaceae and Porphyromonadaceae, were identified to support ginseng's anti-colitis and anti-tumor effects. In addition, alpha and beta diversity data demonstrated that ginseng led to a profound recovery from the AOM/DSS-induced dysbiosis in the microbial community. CONCLUSION: Our results suggest that the CRC chemopreventive effects of American ginseng are mediated through enteric microbiome population-shift recovery and dysbiosis restoration. Ginseng's regulation of the microbiome balance contributes to the maintenance of enteric homeostasis.

Correction to: Role of intestinal microbiome in American ginseng-mediated colon cancer prevention in high fat diet-fed AOM/DSS mice.

This article was originally published with the wrong title.

Purification and properties of neutral beta-N-acetylglucosaminidase from carp blood.

A neutral beta-N-acetylglucosaminidase has been purified to homogeneity from carp blood by a seven-step procedure. It was localized in the cytosol of red blood cells. The purified enzyme was specific to beta-N-acetylglucosaminide and inactive to beta-N-acetylgalactosaminide. It was competitively inhibited by free N-acetylglucosamine, but not by free N-acetylgalactosamine. The optimum pH of the enzyme was 6.5, with a stable pH range of 7.0 to 11.0. The enzyme showed greater heat-lability and anodal electrophoretic mobility than acidic beta-N-acetylglucosaminidases. The Mr value, estimated by sucrose density gradient centrifugation, was 122,000, and the enzyme dissociated into two nonidentical subunits with Mr values of 66,000 and 53,000, based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. With respect to the major characteristics, the neutral enzyme in carp blood was supposed to be a counterpart of hexosaminidase C in human and other mammalian tissues.

Effect of limited proteolysis on the stability and enzymatic activity of human placental S-adenosylhomocysteine hydrolase.

Human placental S-adenosylhomocysteine (AdoHcy) hydrolase was subjected to limited papain digestion. The multiple cleavage sites in the enzyme were identified to be Lys94-Ala95, Tyr100-Ala101, Glu243-Ile244, Met367-Ala368, Gln369-Ile370, and Gly382-Val383. Despite multiple cleavage sites in the backbone of the protein, the digested enzyme was able to maintain its quaternary structure and retain its full catalytic activity. The enzyme activity of the partially digested AdoHcy hydrolase was essentially identical to that of the native enzyme at several pH values. The thermal stabilities of the native and partially digested enzymes were only slightly different at all temperatures tested. The stability of both native and partially digested enzymes were examined in guanidine hydrochloride and equilibrium unfolding transitions were monitored by CD spectroscopy and tryptophan fluorescence spectroscopy. The results of these experiments can be summarized as follows: (1) CD spectroscopic analysis showed that the overall secondary and tertiary structures of the partially digested enzyme are essentially identical with those of the native enzyme; and (2) tryptophan fluorescence spectroscopic analysis indicated that there are small differences in the environments of surface-exposed tryptophan residues between the partially digested enzyme and the native enzyme under unfolding conditions. The differences in the free energy of unfolding, delta(delta Gu) [delta Gu(native)-delta Gu(digested)], is approximately 1.3 kcal/mol. When NAD+ was removed from the partially digested enzyme, the secondary and tertiary structures of the apo form of the digested AdoHcy hydrolase were completely lost and the enzymatic activity could not be recovered by incubation with excess NAD+. These results suggest that AdoHcy hydrolase exists as a very compact enzyme with extensive intramolecular bonding, which contributes significantly to the overall global protein stabilization. Identification of the surface-exposed peptide bonds, which are susceptible to papain digestion, has provided some constraints on the spatial orientations of subunits of the enzyme. This information, in turn, has provided supplemental data for X-ray crystallographic studies currently ongoing in our laboratories.

Methylnaltrexone prevents morphine-induced delay in oral-cecal transit time without affecting analgesia: a double-blind randomized placebo-controlled trial.

Methylnaltrexone is a quaternary opioid antagonist with limited ability to cross the blood-brain barrier and the potential to antagonize the peripherally mediated effects of opioids. The effectiveness of methylnaltrexone in preventing morphine-induced changes in gastrointestinal motility and transit without affecting analgesia was evaluated in humans. Twelve healthy volunteers were given intravenous placebo, placebo plus 0.05 mg/kg morphine, or 0.45 mg/kg methylnaltrexone plus 0.05 mg/kg morphine. Oral-cecal transit time was assessed by the pulmonary hydrogen measurement technique, and analgesia was measured with use of the cold-pressor test. Morphine significantly increased oral-cecal transit time from 104.6 +/- 31.1 minutes (mean +/- SD) to 163.3 +/- 39.8 minutes (p < 0.01). Methylnaltrexone prevented 97% of morphine-induced increase in oral-cecal transit time (106.3 +/- 39.8 minutes; not significant compared with baseline; p < 0.01 compared with morphine alone). Methylnaltrexone did not affect the analgesic effect of morphine on both pain intensity and pain bothersomeness ratings. At a higher dose of morphine (0.1 mg/kg), our preliminary results indicated that 0.45 mg/kg methylnaltrexone also prevented the morphine-induced delay in oral-cecal transit time, with no effect on analgesia. Methylnaltrexone may be a useful adjunct to opioids for the relief of opioid-induced constipation.

The safety and efficacy of oral methylnaltrexone in preventing morphine-induced delay in oral-cecal transit time.

Methylnaltrexone is a quaternary opioid antagonist with limited ability to cross the blood-brain barrier that has the potential to antagonize the peripherally mediated gastrointestinal effects of opioids. In recent trials in human volunteers, we demonstrated that intravenous methylnaltrexone prevented morphine-induced changes in gastrointestinal motility and transit, without affecting analgesia. In this study, 14 healthy volunteers were first given three ascending oral doses of methylnaltrexone to obtain safety and tolerance data (phase A study). In phase B, these subjects were then given single-blind oral placebo and intravenous placebo, followed by randomized, double-blind oral placebo and intravenous morphine (0.05 mg/kg) or oral methylnaltrexone (19.2 mg/kg, an established highest and safe dose based on previous administrations of two smaller doses of 0.64 mg/kg and 6.4 mg/kg in phase A) and intravenous morphine (0.05 mg/kg). Oral-cecal transit time was assessed by the pulmonary hydrogen measurement technique after lactulose ingestion. Morphine significantly increased oral-cecal transit time from 114.6 +/- 37.0 minutes (mean +/- SD) to 158.6 +/- 50.2 minutes (p < 0.001). Oral methylnaltrexone (19.2 mg/kg) completely prevented morphine-induced increase in oral-cecal transit time (110.4 +/- 45.0 minutes; not significant compared with baseline; p < 0.005 compared with morphine alone). These sessions were then followed by single-blind evaluations of descending doses of methylnaltrexone. We observed that 6.4 mg/kg oral methylnaltrexone significantly attenuated the morphine-induced delay in oral-cecal transit time (p < 0.005 compared with morphine alone), and a dose-dependent response was obtained. There was no correlation between oral methylnaltrexone effects on the transit time and the drug plasma concentration, suggesting direct preferential luminal effects of oral methylnaltrexone. Oral methylnaltrexone may have a clinical value in the prevention and treatment of constipation induced by long-term opioid use.

Effects of enteric-coated methylnaltrexone in preventing opioid-induced delay in oral-cecal transit time.

BACKGROUND: Methylnaltrexone is the first peripheral opioid receptor antagonist. It has the potential to prevent or reverse the peripherally mediated gastrointestinal effects of opioids. In previous human volunteer trials, we demonstrated that oral uncoated methylnaltrexone prevented morphine-induced delay in gastrointestinal transit time. METHODS: This trial consisted of two studies: a pilot study and a controlled study. The lactulose hydrogen breath test was used to measure the oral-cecal transit time. RESULTS: In the pilot study with three subjects, an oral dose of 6.4 mg/kg enteric-coated methylnaltrexone effectively reversed the effects of morphine, producing transit times shorter than baseline levels. Subsequently, in the controlled study with another nine subjects, the transit time increased after intravenous morphine administration in all nine subjects, and the lower dose (3.2 mg/kg) of enteric-coated methylnaltrexone completely prevented the morphine-induced change in oral-cecal transit time in all nine subjects. Morphine significantly increased oral-cecal transit time from baseline level of 96.7 +/- 54.1 minutes (mean +/- SD) to 155.0 +/- 53.6 minutes (P = .014). After enteric-coated methylnaltrexone and morphine, the transit time returned to the baseline level (93.3 +/- 56.0 minutes; P = .55 compared with placebo). Plasma concentrations after 6.4 mg/kg and 3.2 mg/kg enteric-coated methylnaltrexone were substantially lower compared with those after 6.4 mg/kg of the uncoated formulation. CONCLUSION: Our results suggest that there is a prevailing direct and local luminal effect of enteric-coated methylnaltrexone and that the enteric-coated formulation exerts its gut pharmacologic actions more efficiently than the uncoated formulation.

Dose-related effects of oral acetaminophen on cold-induced pain: a double-blind, randomized, placebo-controlled trial.

The cold-pressor test is a widely used pain-induction model in humans. This method has been shown to be a sensitive measure for detecting opioid analgesia. However, nonsteroidal anti-inflammatory drugs have not produced consistent analgesic effects with use of this model. The analgesic effect of acetaminophen (INN, paracetamol) on cold pressor-induced pain has not been reported by other investigators. In this study, a double-blind, randomized, placebo-controlled design was used to evaluate the dose-related effects of oral acetaminophen on cold pressor-induced pain in 18 normal healthy human subjects. We observed dose-related analgesic activity of oral acetaminophen using the cold pressor-induced pain model in these subjects. There were statistically significant main effects of both dose and time (pain and bothersomeness ratings decreased with increasing drug dose and increased over time). In pairwise comparisons only the contrast between the highest dose of acetaminophen (1000 mg) and placebo reached statistical significance. Results from our study suggest that the cold-pressor method may have clinical value in evaluating nonopioid analgesic agents.

Gastric effects of methylnaltrexone on mu, kappa, and delta opioid agonists induced brainstem unitary responses.

In this study, we evaluated the gastric effects of methylnaltrexone, an opioid receptor antagonist that does not cross the blood-brain barrier in vivo, on mu, kappa and delta opioid agonists induced brainstem unitary responses in an in vitro neonatal rat brainstem-gastric preparation. Single units in the medial subnucleus of the nucleus tractus solitarius (NTS), responding to electrical stimulation of subdiaphragmatic vagal fibers, were recorded. Selective opioid receptor agonists and antagonists were applied only to the gastric compartment of the bath chamber and thus, the brainstem functions of the preparation were not affected by the drugs. The peripheral gastric effects of a mu opioid receptor agonist, DAMGO, and a kappa opioid receptor agonist, U-50,488H, were evaluated on 58 tonic units that received the subdiaphragmatic vagal inputs. For approximately 78% of the units observed, DAMGO (1.0 microM) and U-50,488H (1.0 microM) induced a concentration-dependent inhibition of 62.1+/-9.3% (mean +/- SE) and 49.2+/-6.5% of the control level of the NTS neuronal activity, respectively. Methylnaltrexone competitively antagonized the DAMGO-induced brainstem neuronal effects. Methylnaltrexone at an 18.8-fold higher concentration also reversed U-50,488H-induced NTS neuronal responses. Naloxone, a non-selective opioid receptor antagonist, reversed the inhibitory effects of DAMGO and U-50,488H at much lower concentrations (3.8% and 0.5%, respectively) compared to methylnaltrexone. Only 18% of the NTS neurons evaluated showed inhibitory responses to a delta receptor agonist, DPDPE, (19.7+/-5.0% at 10 microM), and this inhibition could not be reversed by methylnaltrexone in the concentration range we tested. In addition, when methylnaltrexone (1.0 microM) alone was applied to the gastric compartment, there was an activation (8.5+/-2.1%) of the NTS neurons receiving subdiaphragmatic vagal inputs, suggesting an endogenous gastric opioid action in the modulation of brainstem neuronal activities.

Aristeromycin-5'-carboxaldehyde: a potent inhibitor of S-adenosyl-L-homocysteine hydrolase.

In an earlier study, Liu et al. (Bioorg. Med. Chem. Lett. 1992, 2, 1741-1744) showed that both the E and Z isomers of 4',5'-didehydro-5'-fluoroaristeromycin were very potent irreversible inhibitors of S-adenosylhomocysteine (AdoHcy) hydrolase. However, it was unclear from a mechanistic standpoint whether these vinyl fluorides were themselves type-I mechanism-based inhibitors causing reduction of enzyme-bound NAD+ or whether they were prodrug for aristeromycin-5'-carboxaldehyde, which was the ultimate type-I inhibitor. To elucidate this mechanism of enzyme inhibition, (4'S)- and (4'R)-aristeromycin-5'-carboxaldehydes (1a,b) were synthesized in this study and shown to be potent type-I mechanism-based inhibitors of AdoHcy hydrolase with k2/Ki values of 4.4 x 10(6) and 8.2 x 10(4)M-1min-1, respectively. However, Using 19F NMR and HPLC, it was shown that (4'S)-4,5'-dedehydro-5'-fluoraristeromycin in the presence of AdoHcy hydrolase did not release fluoride ion or generate aristeromycin-5'-carboxaldehyde (1a,b). These results suggest that the E and Z isomers of 4',5'-didehydro-5'-fluoroaristeromycin are inactivating AdoHcy hydrolase by directly reducing NAD+ to NADH and not using the hydrolytic activity of the enzyme to generate aristeromycin-5'-carboxaldehyde.

Inactivation of S-adenosyl-L-homocysteine hydrolase by amide and ester derivatives of adenosine-5'-carboxylic acid.

S-Adenosyl-L-homocysteine (AdoHcy) hydrolase has been shown to have (5'/6') hydrolytic activity with vinyl (5') or homovinyl (6') halides derived from adenosine (Ado). This hydrolytic activity is independent of its 3'-oxidative activity. The vinyl (or homovinyl) halides are converted into 5'(or 6')-carboxaldehydes by the hydrolytic activity of the enzyme, and inactivation occurs via the oxidative activity. Amide and ester derivatives of Ado-5'-carboxylic acid were prepared to further probe the hydrolytic capability of AdoHcy hydrolase. The oxidative activity (but not the hydrolytic activity) is involved in the mechanism of inhibition of the enzyme by the ester and amide derivatives of Ado-5'-carboxylic acid, in contrast to the inactivation of this enzyme by adenosine-derived vinyl or homovinyl halide analogues during which both activities are manifested.

Nucleic acid related compounds. 74. Synthesis and biological activity of 2'(and 3')-deoxy-2'(and 3')-methylenenucleoside analogues that function as mechanism-based inhibitors of S-adenosyl-L-homocysteine hydrolase and/or ribonucleotide reductase.

Treatment of 2-amino-6-chloro-9-(beta-D-ribofuranosyl)purine (21) with TBDMS chloride/imidazole/DMF gave a separable mixture of 5'-O, 2',5'-bis-O (22), 3',5'-bis-O (23), and 2',3',5'-tris-O-TBDMS derivatives. Oxidation of 22 and 23 with CrO3/pyridine/Ac2O, treatment of the respective ketonucleosides with methylenetriphenylphosphorane, and deprotection gave 2-amino-6-chloro-9-[3(and 2)-deoxy-3(and 2)-methylene- beta-D-erythro-pentofuranosyl]purines (28 and 37) that were converted into other 2-amino-6-substituted-purine analogues. Tubercidin was converted into 2'-deoxy-2'-methylenetubercidin (49) by an analogous route. Inactivation of S-adenosyl-L-homocysteine hydrolase by 2'- and 3'-methyleneadenosine analogues was investigated. Mechanism-based inhibition of S-adenosyl-L-homocysteine hydrolase and anticancer and antiviral activities of 2'(and 3')-deoxy-2'(and 3')-methylenenucleoside analogues are discussed.

Anticancer and antiviral effects and inactivation of S-adenosyl-L-homocysteine hydrolase with 5'-carboxaldehydes and oximes synthesized from adenosine and sugar-modified analogues.

Selectively protected adenine nucleosides were converted into 5'-carboxaldehyde analogues by Moffatt oxidation (dimethyl sulfoxide/dicyclohexylcarbodiimide/dichloroacetic acid) or with the Dess-Martin periodinane reagent. Hydrolysis of a 5'-fluoro-5'-S-methyl-5'-thio (alpha-fluoro thioether) arabinosyl derivative also gave the 5'-carboxaldehyde. Treatment of 5'-carboxaldehydes with hydroxylamine [or O-(methyl, ethyl, and benzyl)hydroxylamine] hydrochloride gave E/Z oximes. Treatment of purified oximes with aqueous trifluoroacetic acid and acetone effected trans-oximation to provide clean samples of 5'-carboxaldehydes. Adenosine (Ado)-5'-carboxaldehyde and its 4'-epimer are potent inhibitors of S-adenosyl-L-homocysteine (AdoHcy) hydrolase. They bind efficiently to the enzyme and undergo oxidation at C3' to give 3'-keto analogues with concomitant reduction of the NAD+ cofactor to give an inactive, tightly bound NADH-enzyme complex (type I cofactor-depletion inhibition). Potent type I inhibition was observed with 5'-carboxaldehydes that contain a ribo cis-2',3'-glycol. Their oxime derivatives are "proinhibitors" that undergo enzyme-catalyzed hydrolysis to release the inhibitors at the active site. The 2'-deoxy and 2'-epimeric (arabinosyl) analogues were much weaker inhibitors, and the 3'-deoxy compounds bind very weakly. Ado-5'-carboxaldehyde oxime had potent cytotoxicity in tumor cell lines and was toxic to normal human cells. Analogues had weaker cytotoxic and antiviral potencies, and the 3'-deoxy compounds were essentially devoid of cytotoxic and antiviral activity.