Biomedical subjects
Publications and source records attributed to A Liang.
eRF1a, one of the class-I release factors from ciliate Euplotes octocarinatus, has been crystallized by the vapor-diffusion method using polyethylene glycol 4000 as the precipitant at pH 7.5. The crystal belongs to space group P2(1) and the unit-cell parameters are a=90.4, b=107.9, c=114.8A, beta=94.2 degrees. There appear to be four eRF1a molecules in the asymmetric unit.
Amplification of macronuclear DNA of the ciliate Euplotes octocarinatus revealed the presence of two genes encoding putative polypeptide release factors (RFs) of the codon specific class-I type. They are named eRF1a and eRF1b, respectively. cDNA amplification revealed that both eRF1 genes are expressed. Determination of their copy numbers showed that they are similarly amplified to a level of about 27,000. The deduced protein sequences of the two genes are 57 and 58% identical with human eRF1 and 79% identical to each other. The gene encoding eRF1b possesses three in-frame UGA codons. This codon is known to encode cysteine in Euplotes; only UAA and UAG are used as stop codons in this organism. The primary structure of the two release factors is analyzed and compared with the primary structure of other eukaryotic release factors including the one of Tetrahymena thermophila which uses only UGA as a stop codon. eRF1a and eRF1b of Euplotes as well as eRF1 of Tetrahymena differ from human eRF1 and other class-I release factors of eukaryotes in a domain recently proposed to be responsible for codon recognition. Based on the changes which we observe in this region and the differential use of the stop codons in these two ciliates we predict the amino acids participating in stop codon recognition in eRF1 release factors.
A series of indole and carbazole based inhibitors of factor Xa (FXa) has been investigated. The most potent compound inhibits FXa with a Ki of 0.2 nM and has 900- and 750-fold selectivity over thrombin and trypsin, respectively.
Inhibitors based on the benzimidazole scaffold showed subnanomolar potency against Factor Xa with 500-1000-fold selectivity against thrombin and 50-100-fold selectivity against trypsin. The 2-substituent on the benzimidazole ring had a strong impact on the FXa inhibitory activity. Crystallography studies suggest that the 2-substituent may have a conformational effect favoring the extended binding conformation.
OBJECTIVE: To gain an insight into the hereditary polymorphism of thiopurine methyltransferase(TPMT) activity in Chinese Han population of Shanghai. METHODS: The present authors measured the erythrocyte TPMT activity in 320 healthy Chinese volunteers and 51 children with acute lymphoblastic leukemia(ALL) by means of radiochemical assay. RESULTS: The TPMT activity levels ranged from 4. 32 to 32.33 U/ml pRBCs, with a mean value of (16.64+/-4.50) U/ml pRBCs, male (16.78+/-4.96) U/ml pRBCs, female (16.52+/-4.44) U/ml pRBCs. Eight point one percent of the sample had low activity. The TPMT activity levels for subjects who were <12 years, 13-18 years, 19-45 years and >45 years old were (16.52+/-4.31) U/ml pRBCs, (16. 71+/-4.24) U/ml pRBCs, (16.28+/-5.21) U/ml pRBCs and (17.11+/-3.98) U/ml pRBCs, respectively; the TPMT activity levels for healthy volunteers and patients with ALL were (16.65+/-4.72) U/ml pRBCs and (16.52+/-4.47) U/ml pRBCs, respectively. CONCLUSION: There were no differences of TPMT activity in gender, age, and between healthy volunteers and patients with ALL.
OBJECTIVE: To study the expression of substance P receptor messenger RNA (SP-R mRNA) in nasal mucosa in allergic rhinitis (AR) rat model. METHODS: Twenty Wistar rats free of disease were randomly divided into two groups. AR model rats were established through repeated intraperitoneal shot of ovalbumin (OV) and consequently confirmed by local challenge with OV. SP-R mRNA in nasal mucosa, obtained from two groups, were used to do reverse transcriptive polymerase chain reaction (RT-PCR) and Southern blot. beta-actin was used as a standard control through out the whole process. RESULTS: The results showed definitely that there were positive expression of SP-R mRNA in normal nasal mucosa. This expressions increased significantly (P < 0.05) during nasal allergy. CONCLUSION: Increased expression SP-R mRNA in nasal mucosa in AR model might play roles in the development of AR.
Factor Xa (FXa) is a trypsin-like serine protease that plays a key role in blood coagulation linking the intrinsic and extrinsic pathways to the final common pathway of the coagulation cascade. During our initial studies, we observed facile photochemical conversion of the known FXa/tPA inhibitor, BABCH ¿(E,E)-2, 7-bis(4-amidinobenzylidene)cycloheptan-1-one, 1a, to the corresponding (Z,Z) olefin isomer, 1c (FXa K(i) = 0.66 nM), which was over 25,000 times more potent than the corresponding (E,E) isomer (1a, FXa K(i) = 17 000 nM). In order to determine the scope of this observation, we expanded on our initial investigation through the preparation of the olefin isomers in a homologous series of cycloalkanone rings, 4-substituted cyclohexanone analogues, and modified amidine derivatives. In most cases the order of potency of the olefin isomers was (Z,Z) > (E,Z) > (E,E) with the cycloheptanone analogue (1c) showing the most potent factor Xa inhibitory activity. In addition, we found that selectivity versus thrombin (FIIa) can be dramatically improved by the addition of a carboxylic acid group to the cycloalkanone ring as seen with 8c (FXa K(i) = 6.9 nM, FIIa K(i) > 50,000 nM). Compounds with one or both of the amidine groups substituted with N-alkyl substituents or replaced with amide groups led to a significant loss of activity. In this report we have demonstrated the importance of the two amidine groups, the cycloheptanone ring, and the (Z,Z) olefin configuration for maximum inhibition of FXa within the BABCH template. The results from this study provided the foundation for the discovery of potent, selective, and orally active FXa inhibitors.
A novel series of 2,6-diphenoxypyridines has been designed to inhibit factor Xa, a serine protease strategically located in the coagulation cascade. The evolution from the photochemically unstable bisamidine (Z,Z)-BABCH to potent bisamidine compounds with a pyridine heterocycle as the core scaffold has been achieved. The most potent compound in the series, 6h, has a Ki for human factor Xa of 12 nM. The selectivity of 6h against bovine trypsin and human thrombin was greater than 90- and 1000-fold, respectively. Two proposed modes of binding of 6h to factor Xa are made based on the crystal structures of 6h by itself and of 6h bound to bovine trypsin.
Degradable biomaterials to be used as scaffolds for tissue repair will ideally be able to support new blood vessel growth. The present study evaluated the adherence of human dermal microvascular endothelial cells (HMECs) to an acellular resorbable scaffold material derived from the small intestinal submucosa (SIS). HMECs were exposed to hydrated and dehydrated forms of SIS and to plastic surfaces coated with one of four different known components of the SIS extracellular matrix: collagen Type I, collagen Type IV, fibronectin, and laminin. Results showed that adherence of HMECs to hydrated SIS was greater than to any of the other tested surfaces (P < 0.05). Exposure of HMECs to either soluble collagen Type IV or soluble fibronectin prior to exposure of these cells to hydrated SIS showed only partial inhibition of HMEC attachment. We conclude that HMECs find hydrated SIS to be a suitable substrate for adherence and that dehydration of SIS adversely affects the ability of HMECs to adhere in vitro. The cause of HMEC adherence to SIS appears to be a combination of both its composition and architecture.
A quick and sensitive method has been developed and validated for the determination of tungsten in bulk drug substance and intermediates using either Inductively Coupled Plasma Atomic Emission Spectrometry (ICP-AES) or Inductively Coupled Plasma Mass Spectrometry (ICP-MS). Sample preparation is by direct dissolution with a 80:20 (v/v) concentrated nitric acid:deionized water mixture and avoids labor intensive and potentially hazardous digestion techniques. Excellent agreement was found between ICP-AES and ICP-MS results and between Merck results and Microwave Induced Plasma Mass Spectrometry (MIP-MS) results provided by an independent raw material vendor.
Factor Xa is a serine protease which activates thrombin (factor IIa) and plays a key regulatory role in the blood-coagulation cascade. Factor Xa is, therefore, an important target for the design of anti-thrombotics. Both factor Xa and thrombin share sequence and structural homology with trypsin. As part of a factor Xa inhibitor-design program, a number of factor Xa inhibitors were crystallographically studied complexed to bovine trypsin. The structures of one diaryl benzimidazole, one diaryl carbazole and three diaryloxypyridines are described. All five compounds bind to trypsin in an extended conformation, with an amidinoaryl group in the S1 pocket and a second basic/hydrophobic moiety bound in the S4 pocket. These binding modes all bear a resemblance to the reported binding mode of DX-9065a in bovine trypsin and human factor Xa.
OBJECTIVE: To study in vitro the anticancer interaction between Ys-96, a bisbenzylisoquinoline compound derived from Stephania tetrandra, and adriamycin or vincristine against human cancer cell lines. METHOD: Using human breast cancer cell MCF-7 and its adriamycin-resistant cell line MCF-7/Adr, and human nasopharyngeal cancer cell KB and its vincristine-resistant cell line KBv200 in an in vitro system, anticancer interaction between Ys-96 and adriamycin or vincristine was evaluated with a method reported by Berenbaum. RESULT: The SFIC values (sum of fractional inhibitory concentration) of the combinations with 3 different ratios between Ys-96 and adriamycin or vincristine were markedly less than 1.0, and the shapes of all the isobologram curves were concave. CONCLUSION: The synergistic interaction between Ys-96 and adriamycin or vincristine against the above human cancer cell lines was positively observed in vitro.
OBJECTIVE: To investigate the difference between fresh and dry Dihuang (Radix Rehmanniae) in hemostatic and immunological actions. METHODS: The hemostatic action was assessed in aspirin-treated mice; immunological action was assessed by phagocytosis of macrophages in dexamethasone-treated mice; and Con A-induced mitogenesis and LPS-induced alkaline phosphatase activity of splenocytes were determined in thyroxin-treated mice. RESULT: The blood clotting in mice induced by aspirin was inhibited by oral administrations of juice decoction of fresh Dihuang or decoction of dry Dihuang alike. The action of fresh Dihuang juice was stronger than that of dry Dihuang decoction. Oral administrarion of fresh Dihuang juice or decoction for 10 days enhanced LPS-induced alkaline phosphatase activity of splenocytes in thyroxin-treated mice. The juice also enhanced Con A-induced splenocyte mitogenesis. The action of dry Dihuang decoction was weaker than that of fresh juice in increasing LPS-induced alkaline phosphatase activity. CONCLUSION: It is recommended to follow the tradition of using fresh Dihuang juice or decoction to treat some diseases such as hemorrhagenic troubles.
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The synthesis of amidinoaryloxy 9-benzyl-8-methyl-9H-purine, 7,8-dihydropteridine-6(5H)-one and 5,7-dihydropyrimido[4,5-b][1,4]oxazine-6-one inhibitors of Factor Xa is described. These compounds show nanomolar potency against FXa and maintain high selectivity over thrombin and trypsin.
An arylamidine linker has been employed for the solid-phase synthesis of N-substituted amidinoaryloxypyridine analogs 2 via nucleophilic substitution on a fluoropyridyl template. Two novel N-substituted amidinoaryloxypyridine derivatives 2a and 2b were discovered via this approach.