[Characteristics of different phenotypes of refractory non-erosive gastroesophageal reflux disease].

Objective: To investigate the characteristics of different phenotypes of refractory non-erosive gastroesophageal reflux disease (NERD), the types of esophageal motility and the related factors of symptoms. Methods: A retrospective analysis was made of the patients with refractory NERD of Beijing Tongren Hospital affiliated to Capital Medical University from September 2015 to August 2017. All patients underwent electronic gastroscopy, esophageal manometry and 24-hour dynamic esophageal pH impedance monitoring. They were divided into four phenotype groups according to the results. Results: A total of 231 patients were enrolled in the study. There were 111(48.1%)cases in phenotype 1 group, 9 (3.9%)cases in phenotype 2 group, 100 (43.3%)cases in phenotype 3 group and 11 (4.8%) cases in phenotype 4 group. Compared with the other three groups, the number of weak acid reflux [(86±55) vs (37±8), (70±52), (31±9) times] and the number of gas reflux [(86±76) vs (38±13), (58±57), (26±10)] in phenotype 1 group increased significantly (allP<0.005). Dynamics disorders were common in refractory NERD patients (139/231, 60.2%). Mild esophageal dynamics disorder was the main type of dynamics disorder (118/139, 84.9%). There was no significant difference among the phenotype groups. Multivariate unconditional Logistic regression analysis showed that the risk factors for reflux-related symptoms were female ratio, Chicago power classification, gas reflux and weak acid reflux (OR=3.731, 2.452, 1.036 and 1.037, P<0.05). Conclusions: The characteristics of gastroesophageal reflux and the types of motility disorders are different in different phenotype groups of refractory NERD patients. The risk factors of reflux-related symptoms are female ratio, Chicago motility classification, gas reflux and the frequency of weak acid reflux.

[Related factors of diabetic nephropathy in patients with type 1 diabetes mellitus].

Objective: To analyze the related factors of diabetic nephropathy in inpatients with type 1 diabetes mellitus (T1DM). Methods: A total of 300 patients with T1DM who were treated in the Department of Endocrinology of Anhui Provincial Hospital between 2014 and 2016 were analyzed retrospectively. All the patients were divided into two groups according to their urine albumin-to-creatinine ratio: non-diabetic nephropathy group (n=193) and diabetic nephropathy group (n=107). Multivariate logistic regression analysis was adopted to analyze the factors related to diabetic nephropathy in T1DM, including the age, diabetic duration, body mass index (BMI) and glycosylated hemoglobin (HbA1c) of the two groups. Results: Age, diabetes duration, HbA1c, systolic blood pressure (SBP), diastolic blood pressure (DBP), total cholesterol, triglyceride and uric acid (UA) of diabetic nephropathy group were higher than those of non-diabetic nephropathy group (all P<0.05). The total bilirubin, hemoglobin (Hb) and albumin in non-diabetic nephropathy group were higher than those in diabetic nephropathy group (P<0.05). The proportion of women in diabetic nephropathy group was higher than non-diabetic nephropathy group (62.6% vs 42.0%, P=0.001). Multivariate logistic regression analysis showed that diabetes duration (OR=2.142, 95% CI: 1.011-4.539), HbA1c (OR=1.262, 95% CI: 1.090-1.462), DBP (OR=1.048, 95% CI: 1.001-1.096), UA (OR=1.005, 95% CI: 1.001-1.009) and Hb (OR=0.952, 95% CI: 0.929-0.975) were independent related factors for diabetic nephropathy. Conclusions: Positive controlling of blood pressure, blood glucose, hyperuricemia and correcting anemia may reduce the incidence of diabetic nephropathy in T1DM patients.

[Current diagnosis and treatment progress of small pancreatic neuroendocrine neoplasm].

Small(≤2 cm)pancreatic neuroendocrine neoplasm(pNEN) is a very special subgroup of pNEN featuring a small size, concealed pathogenesis, indolent course and remarkable heterogeneity.Differences in its diagnosis and interventional criteria have evolved from routine pNEN.During recent years, the incidence of small pNEN has increased sharply, while optimal management strategy of this subgroup still remains controversial.In this paper, the biological characteristics, pathological classification, diagnosis, intervention indication and therapeutic principles of small pNEN are reviewed based on recent researches, and current situations of diagnosis and treatment of small pNEN are summarized.

[Clinical useness of multimodal techniques in microsurgical resection of cerebral arteriovenous malformation].

Objective: To explore the clinical useness of intraoperative functional neuronavigation and fluorescent indocyanine green(ICG) angiography as well as electrophysiological evaluation during microsurgical resection of cerebral arteriovenous malformations (AVM). Methods: A series of 42 consecutive cases with AVM underwent microsurgery by intraoperative functional neuronavigation at Department of Neurosurgery of People's Liberation Army General Hospital from January 2009 to February 2015 were retrospectively analyzed. Of the 42 patients, 29 were males and 13 were females aging from 4 to 62 years (mean age 32.6 years). Preoperative assessment included functional magnetic resonance imaging and diffusion tensor imaging to identify the relationship between lesions and eloquent areas. The results of images were integrated into three-dimensional datasets to achieve intraoperative microscopic-based functional neuronavigation during AVM resection. Operations involved in motor areas and corticospinal tract were performed under continuous electrophysiological monitoring. ICG angiography was performed at pre-dissection, post-clipping of the feeders, and post-resection of the nidus. FLOW 800 software presented a color map and ICG intensity-time curve to demostrate the vascular architecture. Postoperative digital subtraction angiography was re-examined routinely to evaluate the extent of resection. Clinical outcomes were evaluated with the modified Rankin Scale. Results: All patients underwent surgery under intraoperative navigation. Of the 42 patients, total resection was achieved in 36 cases (85.7%, 36/42) including 14 cases of AVM in eloquent areas. A total of 40 ICG angiographies were successfully performed among 11 patients. Average number of ICG injections per operation was 3.6 (ranging from 3 to 6). Feeders were visualized in 10 patients and drainers were visualized in 9 cases. The post-surgical follow-up period varied from 3 months to 70 months (mean 22.5 months). 83.8% of the patients returned to normal work and life during the followed-up period. Conclusion: Combining intraoperative neuronavigation and electrophysiological monitoring, as well as fluorescent ICG angiography contribute to microsurgical resection of cerebral AVM effectively in selecting suitable patients, further avoiding neurologic compromise as well.

Low expression of microRNA-21 contributes to LPS-induced osteoblast cell apoptosis through up-regulation of OAS1.

Lipopolysaccharide (LPS) is a critical component of the outer membrane of Gram—negative bacteria. Many cellular signals that are activated by Gram—negative bacteria are initiated by LPS. LPS triggers not only inflammatory responses, but also activates pro—apoptotic signals in a series of human cell types. However, there is relatively minimal data on the microRNA—dependent mechanism(s) of LPS—induced functional activity in osteoblast cells. CCK—8 assay and flow cytometry were used to measure cell viability and apoptosis, respectively. RT—PCR and western blot were performed to determine the mRNA and protein expression in osteoblast cells. In this study, we found that LPS triggered apoptosis in osteoblastic hFOB1.19 cells and induced a low expression of the miRNA—21. Furthermore, through the gene microarray technique, OAS1 was screened and later confirmed to be the target gene which was up—regulated in response to the low expression of miRNA—21. Knockdown of OAS1 by specific siRNAs significantly rescued the LPS—induced hFOB1.19 cell apoptosis. Our data suggest that LPS induces low expression of miRNA—21which consequently causes the up—regulation of the downstream gene OAS1 and eventually triggers apoptosis in hFOB1.19 cells. Knockdown of OAS1 rescues LPS—induced cell death and thus may be a promising therapeutic strategy for orthopedic diseases.

Mesenchymal stem cells transplantation for perianal fistulas: a systematic review and meta-analysis of clinical trials.

BACKGROUND: Perianal fistulas, characterised as granulomatous inflammation of fistulas around the anal canal, are associated with significant morbidity resulting in a negative impact on quality of life and a tremendous burden to the healthcare system. Treatment of anal fistulas usually consists of anal surgery; however, results of closure rates are not satisfactory especially with complex perianal fistulas, after which many patients may suffer from anal incontinence. Recently, the administration of mesenchymal stem cells (MSCs) has shown promising efficacy. Herein, we aim to explore whether MSCs are effective for complex perianal fistulas and if they have either short-term, medium-term, long-term or over-long-term efficacy. Additionally, we want to elucidate whether factors such as drug dosage, MSC source, cell type, and disease aetiology influence treatment efficacy. We searched four online databases and analysed data based on information within the clinical trials registry. The outcomes of eligible trials were analysed with Review Manager 5.4.1. Relative risk and related 95% confidence interval were calculated to compare the effect between the MSCs and control groups. In addition, the Cochrane risk of bias tool was applied to evaluate the bias risk of eligible studies. Meta-analyses showed that therapy with MSCs was superior to conventional treatment for complex perianal fistulas in short-, long- and over-long-term follow-up phases. However, there was no statistical difference in treatment efficacy in the medium term between the two methods. Subgroup meta-analyses showed factors including cell type, cell source and cell dosage were superior compared to the control, but there was no significant difference between different experimental groups of those factors. Besides, local MSCs therapy has shown more promising results for fistulas as a result of Crohn's Disease (CD). Although we tend to maintain that MSCs therapy is effective for cryptoglandular fistulas equally, more studies are needed to confirm this conclusion in the future. SHORT CONCLUSION: MSCs Transplantation could be a new therapeutic method for complex perianal fistulas of both cryptoglandular and CD origin showing high efficacy in the short-term to over-long-term phases, as well as high efficacy in sustained healing. The difference in cell types, cell sources and cell dosages did not influence MSCs' efficacy.

[Clinical effects of anterograde sural neurovascular flap in repairing skin and soft tissue defect around the knee].

Objective: To explore the clinical effects of anterograde sural neurovascular flap in repairing skin and soft tissue defect around the knee. Methods: Nine patients with skin and soft tissue defect around the knee admitted to Beijing Fengtai YouAnMen Hospital from May 2011 to December 2018, were included in this retrospective descriptive study, including 8 males and 1 female, aged 16 to 65 years. The wound area after debridement ranged from 8 cm×5 cm to 18 cm×10 cm. Anterograde sural neurovascular flap was used to repair the wounds in 9 patients, with the area ranging from 9 cm×6 cm to 20 cm×12 cm. The donor sits of flaps in 2 patients were closed and sutured directly, and the donor sits of flaps in 7 patients were repaired with medial split-thickness skin graft of the ipsilateral thigh. The flap survival, complications, and follow-up after operation were recorded. Results: The flaps survived and the blood supply was good in 8 patients and the wounds were closed. One patient developed skin ischemic necrosis which was cured after three weeks of dressing change. All the skin grafts in the donor site of flap in 7 patients survived. In 6 months to 5 years of follow-up after surgery, the skin flap had good texture, color, and shape, and normal sensation. Except for one patient whose knee had poor recovery of function, the knee joint function of the other patients recovered well. Conclusions: The anterograde sural neurovascular flap has the advantages of high survival rate, satisfactory appearance and functional recovery post surgery, and is an ideal flap for repairing the skin and soft tissue defect around the knee.

[Application of bronchoalveolar lavage fluid in patients with pulmonary complications after allogeneic hematopoietic stem cell transplantation].

Objective: To evaluate the diagnostic value of bronchoalveolar lavage (BAL) for pulmonary complications in patients after allogeneic hematopoietic stem cell transplantation (allo-HSCT) and its safety. Methods: Patients with pulmonary complications after allo-HSCT underwent BAL. Microbiological smears, culture, PCR of CMV-DNA, EBV-DNA and TB-DNA, macro genomes new generation sequencing (mNGS) techniques were performed to detect pathogens in BAL fluid (BALF) . Results: A total of 73 allo-HSCT patients with 86 times of pulmonary complications enrolled this prospective study. They underwent 132 times of BAL procedures. The clinical diagnoses of 88.4% cases were made based on BALF analysis. Of them, 67 cases (77.9%) had infectious pulmonary complications, including 29 cases (33.7%) of fungal infection, 18 cases (20.9%) of mixed infection, 11 cases (12.8%) of viral infection and 9 cases (10.5%) of bacterial infection. The other 9 cases (10.5%) of non-infectious pulmonary complications included 8 cases (9.3%) of idiopathic pneumonia syndrome (IPS) and 1 case (1.2%) of pulmonary infiltration of lymphoma. The diagnoses of the remaining 10 cases (11.6%) were not determined. The platelet counts of 33 patients were less than 50×10(9)/L before BAL. None of them developed severe bleeding complications during or after BAL. Transient fever occurred in 10 patients after BAL. Blood cultures showed staphylococcal bacteremia in them and anti-infection therapies were effective. No life-threatening complications occurred in all of the patients during or after BAL. Conclusion: BALF analysis was informative for the diagnosis of pulmonary complication and safe for patients with pulmonary complications after allo-HSCT.

[One case of elderly patient with extremely severe burn complicated by ventricular tachycardia].

One elderly patient with extremely severe burn was admitted to our department on 4th August, 2017. The patient suffered multiple sustained ventricular tachycardia from post injury day 2 to 4 due to relatively high input volume during shock stage. Amiodarone could not be given through intravenous injection because of his low blood pressure. After consultation with cardiologist, ventricular tachycardia was corrected by electrical cardioversion of several times. According to this case, the first treatment is electrical cardioversion when elderly patient with extremely severe burn shows sustained ventricular tachycardia which can not be corrected with medicine.

[Outcomes of reduced-intensity conditioning allogeneic hematopoietic stem cell transplantation for 10 patients with myelofibrosis].

Objective: To evaluate the efficacy of reduced-intensity conditioning allogeneic hematopoietic stem cell transplantation (RIC-allo-HSCT) for patients with myelofibrosis (MF). Methods: The clinical data of 10 patients with myelofibrosis (MF) who underwent RIC-allo-HSCT. Results: Of all 10 patients, 6 were male and 4 women, with a median age of 28.5 (22-54). Using fludarabine/busulfan plus total body irradiation (FB+TBI) pretreatment scheme based. Hematopoiesis reconstitution was achieved in 9 patients (90%). The median time of neutrophil and platelet engraftment was 13.5 (10-22) day and 16.5 (13-40) day, respectively. Acute GVHD occurred in 4 cases while chronic GVHD in 5 cases. The prospective OS for 3 years was (90.0±8.5)% after a median follow-up time of 17 months. Transplant related mortality was 1 case. Conclusion: RIC-HSCT with FB+TBI is a feasible and effective alternative for MF patients.

[Treatment of 30 elderly patients with advanced myeloid neoplasm by allogeneic hematopoietic stem cell transplantation].

Objective: To evaluate the efficacy of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for elderly patients with advanced myeloid neoplasm. Methods: From September 2014 to September 2017, 30 consecutive hospitalized 50-plus-year-old myeloid neoplasm patients were retrospectively analyzed. At the time of transplantation, 6 patients reached complete remission and the others remained no remission after treatment. The donors were identical sibling (12), matched unrelated (6) and haploidentical family member (12), respectively. 18 patients received RIC while 12 patients received MAC conditioning regiments consisted of Busulfan, cytarabine, fludarabine or clarithromycin±TBI, respectively. Results: Five patients died early in the conditioning stage, 24 patients successfully engrafted. The median time of neutrophil engraftment was 14(10-18) d, whereas platelet engraftment was 15(10-19) d. Six cases (25%) experienced aGVHD grades Ⅱ, 8 cases (32%) cGVHD, including moderate to severe cGVHD in 2 cases (8%). Seven, 7 and 5 cases developed CMV viremia, pneumonia and herpeszoster, respectively after transplantation, but no patients died of infections. The median follow-up time of the patients was 7(0.5-38) months. Twenty-one patients were still alive. The estimated 2 years OS and LFS were 62.5% (95% CI 39.2%-85.8%) and 59.2% (95% CI 26.9%-91.5%), respectively. Univariate analysis showed that HCT-CI was the only factor influencing OS. Conclusion: Allogeneic hematopoietic stem cell transplantation could improve the survival of elderly patients with myeloid neoplasm.

Search for new half-metallic ferromagnets in zinc blende CaSi and CaGe by first-principles calculations.

We present an investigation on the electronic structure and ferromagnetism for the hypothetical zinc blende (ZB) CaSi and CaGe by using first-principles full-potential linearized augmented plane-wave (FP-LAPW) calculations. It is found that ZB CaSi and CaGe are half-metallic (HM) ferromagnets without any transition-metal component; they have a magnetic moment of 2.000 µ(B) per formula unit. Analysis of the density of states and magnetic moment indicates that their magnetism mainly originates from the spin polarization of anion p states and the hybridization between the anion p states and the Ca d states. We also find that the half-metallicity can be maintained even when the lattice constant of ZB CaSi and CaGe is compressed up to 8% and 5%, respectively. The absence of the transition-metal atoms makes ZB CaSi and CaGe attractive not only as materials for possible spintronics devices but also as model objects for the study of new mechanisms of the formation of half-metallic ferromagnetism in s-p electron systems.

Anti-thymocyte globulin could improve the outcome of allogeneic hematopoietic stem cell transplantation in patients with highly aggressive T-cell tumors.

The early experiment result in our hospital showed that anti-thymocyte globulin (ATG) inhibited the proliferation of lymphoid tumor cells in the T-cell tumors. We used the ATG as the part of the conditioning regimen and to evaluate the long-term anti-leukemia effect, the safety and complication in the patients with highly aggressive T-cell lymphomas. Twenty-three patients were enrolled into this study. At the time of transplant, six patients reached first or subsequent complete response, three patients had a partial remission and 14 patients had relapsed or primary refractory disease. The conditioning regimen consisted of ATG, total body irradiation, toposide and cyclophosphamide. The complete remission rate after transplant was 95.7%. At a median follow-up time of 25 months, 16 (69.6%) patients are alive and free from diseases, including nine patients in refractory and progressive disease. Seven patients died after transplant, five from relapse and two from treatment-related complications. The incidence of grades II-IV acute graft-vs-host disease (GvHD) was 39.1%. The maximum cumulative incidence of chronic GvHD was 30%. The most frequent and severe conditioning-related toxicities observed in 8 out of 23 patients were grades III/IV infections during cytopenia. Thus, ATG-based conditioning is a feasible and effective alternative for patients with highly aggressive T-cell tumors.

Pharmacological characterization of novel A3 adenosine receptor-selective antagonists.

The effects of putative A3 adenosine receptor antagonists of three diverse chemical classes (the flavonoid MRS 1067, the 6-phenyl-1,4-dihydropyridines MRS 1097 and MRS 1191, and the triazoloquinazoline MRS 1220) were characterized in receptor binding and functional assays. MRS1067, MRS 1191 and MRS 1220 were found to be competitive in saturation binding studies using the agonist radioligand [125I]AB-MECA (N6-(4-amino-3-iodobenzyl)adenosine-5'-N-methyluronamide) at cloned human brain A3 receptors expressed in HEK-293 cells. Antagonism was demonstrated in functional assays consisting of agonist-induced inhibition of adenylate cyclase and the stimulation of binding of [35S]guanosine 5'-O-(3-thiotriphosphate) ([35S]GTP-gamma-S) to the associated G-proteins. MRS 1220 and MRS 1191, with KB values of 1.7 and 92 nM, respectively, proved to be highly selective for human A3 receptor vs human A1 receptor-mediated effects on adenylate cyclase. In addition, MRS 1220 reversed the effect of A3 agonist-elicited inhibition of tumor necrosis factor-alpha formation in the human macrophage U-937 cell line, with an IC50 value of 0.3 microM.

6-phenyl-1,4-dihydropyridine derivatives as potent and selective A3 adenosine receptor antagonists.

An approach to designing dihydropyridines that bind to adenosine receptors without binding to L-type calcium channels has been described. 1,4-Dihydropyridine derivatives substituted with beta-styryl or phenylethynyl groups at the 4-position and aryl groups at the 6-position were synthesized and found to be selective for human A3 receptors. Combinations of methyl, ethyl, and benzyl esters were included at the 3- and 5-positions. Affinity was determined in radioligand binding assays at rat brain A1 and A2A receptors using [3H]-(R)-PIA [[3H]-(R)-N6-(phenylisopropyl)adenosine] and [3H]CGS 21680 [[3H]-2-[[4-(2-carboxyethyl)phenyl]ethylamino]-5'- (N-ethylcarbamoyl)adenosine], respectively. Affinity was determined at cloned human and rat A3 receptors using [125I]AB-MECA [N6-(4-amino-3-iodobenzyl)-5'-(N-ethylcarbamoyl)-adenosine]. Structure-activity analysis indicated that substitution of the phenyl ring of the beta-styryl group but not of the 6-phenyl substituent was tolerated in A3 receptor selective agents. Replacement of the 6-phenyl ring with a 3-thienyl or 3-furyl group reduced the affinity at A3 receptors by 4- and 9-fold, respectively. A 5-benzyl ester 4-trans-beta-styryl derivative, 26, with a Ki value of 58.3 nM at A3 receptors, was > 1700-fold selective vs either A1 receptors or A2A receptors. Shifting the benzyl ester to the 3-position lowered the affinity at A3 receptors 3-fold. A 5-benzyl, 3-ethyl ester 4-phenylethynyl derivative, 28, displayed a Ki value of 31.4 nM at A3 receptors and 1300-fold selectivity vs A1 receptors. The isomeric 3-benzyl, 5-ethyl diester was > 600-fold selective for A3 receptors. Oxidation of 28 to the corresponding pyridine derivative reduced affinity at A3 receptors by 88-fold and slightly increased affinity at A1 receptors.

Synthesis and HIV inhibition activity of 2',3'-dideoxy-3'-C-hydroxymethyl nucleosides.

A series if 2',3'-dideoxy-3'-C-hydroxymethyl purine nucleosides were prepared based on the photochemical ring expansion of a chiral cyclobutanone precursor, (2S)-trans-2,3-bis[(benzoyloxy)methyl]cyclobutanone, in the presence of a 6-substituted purine. Both alpha- and beta-anomers are produced in this transformation. Deprotection was effected by reaction of the photoadducts with saturated methanolic ammonia. Nine purine nucleosides were tested for their inhibitory effect of HIV IIIB virus on H9 cells. The 6-hexyloxy and adenine derivatives 4e,c, respectively, appeared to be most effective at inhibiting viral reproduction with 4c comparable in activity to ddI and AZT.

Synthesis and biological activities of flavonoid derivatives as A3 adenosine receptor antagonists.

A broad screening of phytochemicals has demonstrated that certain flavone and flavonol derivatives have a relatively high affinity at A3 adenosine receptors, with Ki values of > or = 1 microM (Ji et al. J. Med. Chem. 1996, 39, 781-788). We have further modified the flavone structure to achieve a degree of selectivity for cloned human brain A3 receptors, determined in competitive binding assays versus [125I]AB-MECA[N6-(4-amino-3-iodobenzyl)adenosine-5'-(N-methylur onamide)]. Affinity was determined in radioligand binding assays at rat brain A1 and A2a receptors using [3H]-N6-PIA ([3H]-(R)-N6-phenylisopropyladenosine) and [3H]CGS21680 [[3H]-2-[[4-(2-carboxyethyl)phenyl]ethylamino]-5'-(N-ethylcarbamoyl++ +)adenosine], respectively. The triethyl and tripropyl ether derivatives of the flavonol galangin, 4, had Ki values of 0.3 - 0.4 microM at human A3 receptors. The presence of a 5-hydroxyl group increased selectivity of flavonols for human A3 receptors. The 2',3,4',7-tetraethyl ether derivative of the flavonol morin, 7, displayed a Ki value of 4.8 microM at human A3 receptors and was inactive at rat A1/A2a receptors. 3,6-Dichloro-2'-(isopropyloxy)-4'-methylflavone, 11e, was both potent and highly selective (approximately 200-fold) for human A3 receptors (Ki = 0.56 microM). Among dihydroflavonol analogues, the 2-styryl instead of the 2-aryl substituent, in 15, afforded selectivity for human A3 vs rat A1 or A2A receptors. The 2-styryl-6-propoxy derivative, 20, of the furanochromone visnagin was 30-fold selective for human A3 receptors vs either rat A1 or A2A receptors. Several of the more potent derivatives effectively antagonized the effects of an agonist in a functional A3 receptor assay, i.e. inhibition of adenylyl cyclase in CHO cells expressing cloned rat A3 receptors. In conclusion, these series of flavonoids provide leads for the development of novel potent and subtype selective A3 antagonists.

Interaction of 1,4-dihydropyridine and pyridine derivatives with adenosine receptors: selectivity for A3 receptors.

1,4-Dihydropyridine and pyridine derivatives bound to three subtypes of adenosine receptors in the micromolar range. Affinity was determined in radioligand binding assays at rat brain A1 and A2A receptors using [3H]-(R)-PIA [[3H]-(R)-N6-(phenylisopropyl)adenosine] and [3H]CGS 21680 [[3H]-2-[[4-(2-carboxyethyl)phenyl]ethylamino]-5'-(N-ethylcarbamoyl++ +) adenosine], respectively. Affinity was determined at cloned human and rat A3 receptors using [125I]AB-MECA [N6-(4-amino-3-iodobenzyl)-5'-(N-methylcarbamoyl)adenosine]. Structure-activity analysis at adenosine receptors indicated that sterically bulky groups at the 4-, 5-, and 6-positions are tolerated. (R,S)-Nicardipine, 12, displayed Ki values of 19.6 and 63.8 microM at rat A1 and A2A receptors, respectively, and 3.25 microM at human A3 receptors. Similarly, (R)-niguldipine, 14, displayed Ki values of 41.3 and 1.90 microM at A1 and A3 receptors, respectively, and was inactive at A2A receptors. A preference for the R- vs the S-enantiomer was observed for several dihydropyridines at adenosine receptors, in contrast with the selectivity at L-type Ca2+ channels. A 4-trans-beta-styryl derivative, 24, with a Ki value of 0.670 microM at A3 receptors, was 24-fold selective vs A1 receptors (Ki = 16.1 microM) and 74-fold vs A2A receptors (Ki = 49.3 microM). The affinity of 24 at L-type Ca2+ channels, measured in rat brain membranes using [3H]isradipine, indicated a Ki value of 0.694 microM, and the compound is thus nonselective between A3 receptors and L-type Ca2+ channels. Inclusion of a 6-phenyl group enhanced A3 receptor selectivity: Compound 28 (MRS1097; 3,5-diethyl 2-methyl-6-phenyl-4-(trans-2-phenylvinyl)-1,4(R,S)-dihydro-pyridin e-3, 5-dicarboxylate) was 55-fold selective vs A1 receptors, 44-fold selective vs A2A receptors, and over 1000-fold selective vs L-type Ca2+ channels. In addition, compound 28 attenuated the A3 agonist-elicited inhibitory effect on adenylyl cyclase. Furthermore, whereas nicardipine, 12, displaced radioligand from the Na(+)-independent adenosine transporter with an apparent affinity of 5.36 +/- 1.51 microM, compound 28 displaced less than 10% of total binding at a concentration of 100 microM. Pyridine derivatives, when bearing a 4-alkyl but not a 4-phenyl group, maintained affinity for adenosine receptors. These findings indicate that the dihydropyridines may provide leads for the development of novel, selective A3 adenosine antagonists.

D-penicillamine induces rat hepatic metallothionein.

Intraperitoneal injection of D-Penicillamine (D-PA) at a dose range of 20-500 mg/kg increased rat hepatic but not renal and pancreatic metallothionein (MT). Elevated MT predominantly contained Zn. Maximal induction was obtained 18 h after a single injection of 200 mg D-PA/kg resulting in 148 +/- 18 micrograms MT/g liver which was 16.4-times the control level of 9 +/- 2 micrograms MT/g. At 48 h after injection, MT declined to 18 +/- 9 micrograms MT/g liver. At maximal MT increase the content of total hepatic Zn but not of Cu was elevated. Increased amounts of Zn in liver homogenate, cytosol and MT could be detected approximately 4 h after injection of 200 mg D-PA/kg. Concomitantly there was a decrease in Zn bound to cytosolic non-MT ligands. All Zn changes reversed at 18 h. These data show that already single doses of D-PA cause induction of Zn-thionein in rat liver and lead to synchronous redistribution of Zn from endogenous sources to newly synthesized MT.