[Reliability and validity of the Chinese version of the test of the adherence to inhalers (TAI)].

Objectives: To evaluate the reliability and validity of the Chinese version of the test of the adherence to inhalers (TAI) in Chinese patients with chronic airway disease. Methods: Based on the English version of TAI, the items of the Chinese version of TAI were determined after forward-backward translation and cultural adaption. Totally, 165 patients with chronic obstructive pulmonary disease (COPD) and asthma were enrolled from Respiratory Clinic of the Second Xiangya Hospital of Central South University from July to November 2021, and a questionnaire survey was conducted using the Chinese version of TAI and the Morisky medication adherence scale 8-item version (MMAS-8). The content validity of the scale was expressed by content validity index (CVI) and the construct validity was analyzed by exploratory factor analysis (EFA). The convergence validity was evaluated by Pearson correlation analysis. The reliability of the scale was expressed by Cronbach's α coefficient, the split-half reliability and test-retest reliability. Results: The CVI was 0.966. There were 10 items in total. Two factors were extracted from the Chinese version of TAI and the cumulative variance contribution rate was 57.236%. The load value of each item was more than 0.400 and the factor attribution of the item was consistent with the original scale. The total score of the Chinese version of TAI was positively correlated with the total score of the MMAS-8(r=0.835,P<0.001). The Cronbach's α of the overall scale was 0.843, the Guttman's half-reliability coefficient was 0.796 and the test-retest reliability was 0.884 (P<0.001), respectively. Conclusions: The Chinese version of TAI has good reliability and validity, which may be a reliable tool for evaluating the adherence to inhalers of patients with chronic airway disease in China.

Spironolactone prevents dietary-induced metabolic syndrome by inhibiting PI3-K/Akt and p38MAPK signaling pathways.

OBJECTIVES: Aim of the study is to evaluate the impact of spironolactone (SPL) on indexes of metabolic syndrome (MS) and further investigate the mechanisms underlying its protective effects. METHODS: A rat model of MS was established by administering a fat- and salt-enriched diet (FS diet). The occurrence of MS was examined by measurement of blood pressure (BP), aldosterone (ALD) content, blood lipid (BL), glucose and insulin levels. Homeostasis model assessment of insulin resistance (HOMA-IR) was calculated. Pancreatic gland tissue injury was assessed by ß-cell apoptosis. Mineralocorticoid receptor (MR) activity, phosphatidylinositol 3- kinase/Akt (PI3-K/Akt), and phosphorylation of p38MAPK (Pp38MAPK) in pancreatic gland tissue were evaluated by western blot analysis. RESULTS: SPL prevented hypertension, and dyslipidemia during MS induced by the intake of FS diet, but had no effect on K+ and Na+ disturbances. Furthermore, SPL significantly attenuated ALD and MR expression levels after FS diet. Finally, SPL inhibited phosphorylation protein kinase B (p- PKB) activation in the pancreatic gland tissue, a downstream target of PI3-K, and phosphorylation of p38MAPK pathway, critical for cellular apoptosis. CONCLUSIONS: This study demonstrates that SPL exerts a protective effect on hypertension and dyslipidemia. This protective effect may depend, at least in part, on MAPK and PI3-K pathways.

[Relative bioavailability of effervescent tablet of potassium chloride in healthy volunteers].

AIM: To probe the approach by which the pharmacokinetics and relative bioavailability of endogenous medicinal substances can be studied. METHODS: A randomized three-crossover study was performed in 18 healthy male volunteers. In two of the three study periods, a single 2 g dose of either effervescent tablet or common tablet of potassium chloride was administered; whereas in one of three periods no drug treatment was given to allow the nondrug-related (endogenous) potassium in urine to be determined. In each period the urine samples were collected at the following intervals: 0-2, 2-4, 4-6, 6-8, 8-10, 10-12, 12-24, 24-48 h after dose. Urine potassium was determined and the cumulative urine potassium-time data were fitted to a one-compartment model with first-order absorption. Bioavailability was represented by cumulative amount of potassium excreted in urine during 48 hours after drug administration and the bioequivalence of the two formulations was evaluated by analysis of variance and two one-sided t-test. RESULTS: The pharmacokinetic parameters were as follows: effervescent tablet T1/2 ke = (6 +/- 5) h, T1/2 ka = (0.08 +/- 0.08) h, ku = (0.09 +/- 0.04) h-1, Xmax/f = (18 +/- 8) mmol; common tablet T1/2 ke = (8 +/- 5) h, T1/2 ka = (0.11 +/- 0.11) h, ku = (0.07 +/- 0.04) h-1, Xmax/f = (18 +/- 8) mmol. Relative bioavailability of effervescent tablet was 97.5% +/- 15.2% compared with common tablet. CONCLUSION: The two formulations were of bioequivalence. The methods used in this study might be applicable to other similar studies involving endogenous medicinal substances.

Conformational correlation and coupled motion between residue A21 and B25 side chain observed in crystal structures of insulin mutants at position A21.

The C-terminal residue of the insulin A chain is invariant and kept as asparagine in all known insulin molecules from hagfish through birds to mammals. To get information on the role of this conserved residue, which is still unclear, the three-dimensional structures of four human insulin mutants, A21 Asn-->Gly, A21 Asn-->Asp, A21 Asn-->Ala, and A21 Asn-->Gln DesB30, were determined by X-ray crystallography. The four mutants crystallize separately into two kinds (rhombohedral and cubic) of crystals. In the refined structures, conformational correlation and coupled motion between the A chain C-terminal residue A21 and the B25 side chain was observed, in contrast to the nearly unchanged general structures as compared with the native insulin structures in their respective crystals. A detailed analysis suggests that residue A21 can affect insulin receptor binding by interaction with the B25 side chain and the B chain C-terminal segment to assist the B25 side chain rearranging into the 'active' conformation.

Crystal structures of two alpha-like scorpion toxins: non-proline cis peptide bonds and implications for new binding site selectivity on the sodium channel.

The crystal structures of two group III alpha-like toxins from the scorpion Buthus martensii Karsch, BmK M1 and BmK M4, were determined at 1.7 A and 1.3 A resolution and refined to R factors of 0.169 and 0.166, respectively. The first high-resolution structures of the alpha-like scorpion toxin show some striking features compared with structures of the "classical" alpha-toxin. Firstly, a non-proline cis peptide bond between residues 9 and 10 unusually occurs in the five-member reverse turn 8-12. Secondly, the cis peptide 9-10 mediates the spatial relationship between the turn 8-12 and the C-terminal stretch 58-64 through a pair of main-chain hydrogen bonds between residues 10 and 64 to form a unique tertiary arrangement which features the special orientation of the terminal residues 62-64. Finally, in consequence of the peculiar orientation of the C-terminal residues, the functional groups of Arg58, which are crucial for the toxin-receptor interaction, are exposed and accessible in BmK M1 and M4 rather than buried as in the classical alpha-toxins. Sequence alignment and characteristics analysis suggested that the above structural features observed in BmK M1 and M4 occur in all group III alpha-like toxins. Recently, some group III alpha-like toxins were demonstrated to occupy a receptor site different from the classical alpha-toxin. Therefore, the distinct structural features of BmK M1 and M4 presented here may provide the structural basis for the newly recognized toxin-receptor binding site selectivity. Besides, the non-proline cis peptide bonds found in these two structures play a role in the formation of the structural characteristics and in keeping accurate positions of the functionally crucial residues. This manifested a way to achieve high levels of molecular specificity and atomic precision through the strained backbone geometry.

Structure of an insulin dimer in an orthorhombic crystal: the structure analysis of a human insulin mutant (B9 Ser-->Glu).

The structure of human insulin mutant B9 (Ser-->Glu) was determined by an X-ray crystallographic method at 2.5 A resolution with an R factor of 0.165 under non-crystallographic restraints. The crystals were grown at low pH (<3.8) and belong to the orthorhombic P2(1)2(1)2(1) space group with unit-cell dimensions a = 44.54, b = 46.40, c = 51.85 A and one dimer per asymmetric unit without further aggregation. The structure in this crystal form can be regarded as a model for a discrete insulin dimer and displays the following features compared with the structure of 2Zn insulin. (i) The overall dimer is expanded and more symmetric. The two A chains are about 2 A more distant from each other, while the two B chains are about 0.8 A further apart. Both monomers are more similar to molecule 1 than molecule 2 of the 2Zn insulin dimer. (ii) The dimer structure is stabilized by protonation and neutralization of the carboxyl groups at lower pH and, in addition, by formation of a hydrogen-bond network among the side chains of residues GluB9, HisB13 and HisB10 on the dimer-forming surface of both monomers, resulting from a structural rearrangement. (iii) The B-chain amino-terminal segment is in an open state (O state), i.e. a state different from the well known R and T states found in the insulin hexamer. In the O state, the B-chain N-terminal segment is in an extended conformation and is detached from the rest of the molecule. This conformational state has also been observed in the monomeric crystal structure of despentapeptide (B26-B30) and desheptapeptide (B24-B30) insulin, as well as in the solution structure of an engineered insulin monomer. It suggests that the O state may be the characteristic conformation of insulin in lower aggregation forms and may be relevant to the formation of insulin fibrils. In addition, based on the crystallization process, the smallest possible building blocks of insulin crystal are also discussed.

Crystallization and preliminary crystallographic analyses of pokeweed antiviral protein from seeds.

Pokeweed antiviral protein from seeds (PAP-S) is a ribosome inactivating protein which has lowest toxicity and highest inhibition activity as opposed to other pokeweed antiviral proteins and its three potential glycosylation sites (10, 44, 255) were shown to bind to N-acetylglucosamine. Good quality crystals of PAP-S were grown at high protein concentration (100 mg ml-1) and high temperature (306 K). The crystals have space group I222 and cell parameters a = 78.7, b = 85.2 and c = 93.0 A. An X-ray diffraction data set with resolution up to 1.8 A was collected. This high-resolution data will help to locate the sugars bound to the protein and provide accurate structural data for understanding structure-function relationships of PAP-S.

[The detection and study on enterotoxigenic Escherichia coli in China].

In this paper, some strains of ETEC were first detected from the stools of the patients with cholerae-like diarrhea in China in 1974. With a detected rate 36.48% of ETEC Detected rates weve 20.18% from patients with acute diarrhea, 19.74% from environment and sea foods. ETEC produced most of LT toxin in culture medium when the they were incubated at 37-42 degrees C for 4 days. When Rabbit ileal loop test (RILT) was performed with the culture of ETEC or with germ-free culture supernatant, a large quantity of intestinal fluid in response to LT was accumulated at 24 hour and showed positive reaction at every test. Similar results were qained by using 5 cm or 10 cm ileal loop of RILT for LT detectoing. Better effects were noticed through injection of 1.0-3.0 ml culture of ETEC or germ-free enterotoxin in per 5 cm ileal loop. LT could be destroyed by heating at > 50 degrees C or pH < 4.0.

Crystal structure of an acidic neurotoxin from scorpion Buthus martensii Karsch at 1.85 A resolution.

The crystal structure of an acidic scorpion neurotoxin, BmK M8, purified from Chinese scorpion Buthus martensii Karsch (BmK), has been determined by the molecular replacement method. It is the first structure of an acidic alpha-scorpion neurotoxin reported so far. The crystals adopt a symmetry of space group P2(1) and contain one molecule per asymmetric unit. The structure has been refined to an R factor of 18.1% using reflection data in the range of 8 to 1.85 A resolution, with standard deviations from ideal geometry of 0.017 A and 2.43 degrees for bond length and angle, respectively. The 12 residues at the C terminus with unknown sequence were determined by crystallographic refinement. The refined model shows that the structural core, consisting of a motif beta alpha beta beta, is similar to that of toxin II from Androctonus australis Hector (AaH II) or Variant 3 from Centruroides sculpturatus Ewing (CsE V3). The three conformationally variable loops protruding from this structural core are different from that of AaH II, and especially from that of CsE V3. Compared with the most potent and basic alpha-toxin AaH II, the BmK M8 is a relatively inactive toxin (1100 times less active than AaH II) with an unusually low isoelectric point (pI 5.3). Sequence alignment of the two toxins shows a difference of 26 residues (40.6%). Among them four basic or neutral residues in AaH II, namely Val10, Lys28, Val55 and Gly59, are changed to acidic glutamate in BmK M8. The residues Glu10, Glu28 and Glu55 of BmK M8 are located on a surface (Face B), opposite the "conserved hydrophobic surface" (Face A). The latter is a functionally important area proposed by Fontecilla-Camps et al. Our observations suggest that in addition to Face A, Face B may also be involved in the biological activity of scorpion toxins. The structure of BmK M8 shows an evident conformational change of the alpha-amino group at the N terminus and a deorganization of Arg2 caused by the mutation D53A. These structural changes may also be responsible for the weak toxicity of BmK M8. In association with the information from chemical modifications, a multisite binding mode for toxin-receptor interaction and three "toxic regions" in relevance to the binding process, including Face A, Face B and Site C, are proposed. Face A, mainly consisting of Tyr5, 35, 47, the alpha-amino group, Arg2 and Asp3, may be more essential for the binding. Face B, mainly comprising conserved residues Tyr14, 21, Lys28 and Val55, may contribute to the high efficacy of the binding process and substitutions by acidic residues in this area could strongly weaken the toxic activity. Site C, formed by Lys58 and Arg62 at the C terminus and Arg41 and Tyr42 from loop 38-44, may be involved in binding site specificity.

[Studies on the components of trace elements and macro elements in hemolymph of Anopheles anthropophagus].

The elements in hemolymph of An. anthropophagus were determined by ICAP. There were 18 kinds of trace elements such as Fe,Zn,Cu,Mn,Cr,Mo,Co,Ni,V,Sr,B,Al,Ba,Zr, Cd,Pb,Ga,Li and 6 kinds of macro elements such as Ca,Mg,K,Na,S,P in the hemolymph of the mosquito. The contents of the macro elements and Fe in hemolymph of newly emerged mosquitoes were significantly higher than those of mosquitoes after taking blood meal, whereas Zn and Al were lower. Comparing elements in hemolymph of An. anthropophagus and An. sinensis, there were 14 kinds of elements in newly emerged mosquitoes with striking significant difference, while there were 13 kinds of elements with striking significant difference in the mosquitoes after taking blood meal. Comparing elements in the hemolymph of An. anthropophagus and Ae. albopictus, there were 13 kinds of elements with striking significant difference in the hemolymph of newly emerged mosquitoes and the mosquitoes after taking blood meal. The results suggested that the components of elements in hemolymph were relevant to the nutritional metabolism and development of mosquitoes, and that mosquitoes of different species and with different sensibilities to malaria parasites also showed difference in the contents of elements in their hemolymph.