Crystal structure of β-hexosaminidase B in complex with pyrimethamine, a potential pharmacological chaperone

β-Hexosaminidases (β-hex) are a group of glycosyl hydrolase isozymes that break down neutral and sialylated glycosphingolipids in the lysosomes, thereby preventing their buildup in neuronal cells. Some mutants of β-hex have decreased folding stability that results in adult-onset forms of lysosomal storage diseases. However, prevention of the harmful accumulation of glycolipids only requires 10% of wild-type activity. Pyrimethamine (PYR) is a potential pharmacological chaperone that works by stabilizing these mutant enzymes sufficiently to allow more β-hex to arrive in the lysosome, where it can carry out its function. An X-ray structure of the complex between human β-hexosaminidase B (HexB) and PYR has been determined to 2.8 ?. PYR binds to the active site of HexB where several favorable van der Waals contacts and hydrogen bonds are introduced. Small adjustments of the enzyme structure are required to accommodate the ligand, and details of the inhibition and stabilization properties of PYR are discussed.     

Special issue on molecular targets, biomarkers and animal models for anti-cancer pharmacological research： potentials and challenges from chemoprevention to chemotherapeutic treatment

With the current advances in the field of cancer research particularly in cancer chemoprevention, we have much to celebrate but we also have a lot of challenges before us. Some of the challenges include finding appropriate animal models for predicting efficacy of chemopreventive compounds for human cancers, of which are usually highly complex and involved multiple genes in nature and translating our basic research into clinical practice. One of the other challenges will be to disseminate our ideas into the larger biomedical and pharmaceutical community. With this in mind, I have been asked by the Editorial Team of Acta Pharmacologica Sinica to organize a special issue to discuss the current advances in cancer chemoprevention from prevention to treatment. The distinction and the potential overlapping nature of cancer chemoprevention versus chemotherapeutic treatment has become somewhat blurred, since prevention of carcinogenesis encompasses preventing any or all of the three stages of carcinogenesis namely initiation, promotion and progression. Oftentimes, many cancer chemopreventive compounds can possess all three biological properties that culminate in the overall protection against cancer. To be most effective, cancer chemoprevention of earlier lesions will be the ultimate goal if we are to eradicate this dreadful disease, since advanced metastasized cancers are almost lethal and unresponsive to radiation and chemotherapy. The reviews covered in this special issue, which are based on strong preclinical studies, will hopefully provide the impetus to translate this information to clinical practice and improved patient care.     

Interaction of hydrogen sulfide with ion channels

1. Hydrogen sulfide (H₂S) is a signalling gasotransmitter. It targets different ion channels and receptors, and fulfils its various roles in modulating the functions of different systems. However, the interaction of H₂S with different types of ion channels and underlying molecular mechanisms has not been reviewed systematically. 2. H₂S is the first identified endogenous gaseous opener of ATP‐sensitive K⁺ channels in vascular smooth muscle cells. Through the activation of ATP‐sensitive K⁺ channels, H₂S lowers blood pressure, protects the heart from ischemia and reperfusion injury, inhibits insulin secretion in pancreatic β cells, and exerts anti‐inflammatory, anti‐nociceptive and anti‐apoptotic effects. 3. H₂S inhibited L‐type Ca²⁺ channels in cardiomyocytes but stimulated the same channels in neurons, thus regulating intracellular Ca²⁺ levels. H₂S activated small and medium conductance K_Ca channels but its effect on BK_Ca channels has not been consistent. 4. H₂S‐induced hyperalgesia and pro‐nociception seems to be related to the sensitization of both T‐type Ca²⁺ channels and TRPV₁ channels. The activation of TRPV₁ and TRPA₁ by H₂S is believed to result in contraction of nonvascular smooth muscles and increased colonic mucosal Cl^? secretion. 5. The activation of Cl^? channel by H₂S has been shown as a protective mechanism for neurons from oxytosis. H₂S also potentiates N‐methyl‐d ‐aspartic acid receptor‐mediated currents that are involved in regulating synaptic plasticity for learning and memory. 6. Given the important modulatory effects of H₂S on different ion channels, many cellular functions and disease conditions related to homeostatic control of ion fluxes across cell membrane should be re‐evaluated.     

Hydrogen sulfide facilitates carotid sinus baroreflex in anesthetized rats

AIM: To study effects of hydrogen sulfide (H2S) on the carotid sinus baroreflex (CSB). METHODS: The functional curve of the carotid sinus baroreflex was measured by recording changes in arterial pressure in anesthetized male rats with perfused carotid sinus. RESULTS: H2S (derived from sodium hydrosulfide) at concentrations of 25, 50, and 100 micromol/L facilitated the CSB, shifting the functional curve of the baroreflex downward and to the left. There was a marked increase in peak slope (PS) and reflex decrease in blood pressure (RD). Effects were concentration-dependent. Pretreatment with glibenclamide (20 micromol/L), a K(ATP) channel blocker, abolished the above effects of H2S on CSB. Pretreatment with Bay K8644 (an agonist of calcium channels; 500 nmol/L) eliminated the effect of H2S on CSB. An inhibitor of cystathionine gamma-lyase (CSE), DL-propargylglycine (PPG; 200 micromol/L), inhibited CSB in male rats and shifted the functional curve of the baroreflex upward and to the right. CONCLUSION: These data suggest that exogenous H2S exerts a facilitatory role on isolated CSB through opening K(ATP) channels and further closing the calcium channels in vascular smooth muscle. Endogenous H2S may activate the activity of the CSB in vivo.     

Cytochrome oxidase inhibition induced by acute hydrogen sulfide inhalation: correlation with tissue sulfide concentrations in the rat brain, liver, lung, and nasal epithelium.

Hydrogen sulfide (H2S) is an important brain, lung, and nose toxicant. Inhibition of cytochrome oxidase is the primary biochemical effect associated with lethal H2S exposure. The objective of this study was to evaluate the relationship between the concentration of sulfide and cytochrome oxidase activity in target tissues following acute exposure to sublethal concentrations of inhaled H2S. Hindbrain, lung, liver, and nasal (olfactory and respiratory epithelial) cytochrome oxidase activity and sulfide concentrations were determined in adult male CD rats immediately after a 3-h exposure to H2S (10, 30, 80, 200, and 400 ppm). We also determined lung sulfide and sulfide metabolite concentrations at 0, 1.5, 3, 3.25, 3.5, 4, 5, and 7 h after the start of a 3-h H2S exposure to 400 ppm. Lung sulfide concentrations increased during H2S exposure and rapidly returned to endogenous levels within 15 min after the cessation of the 400-ppm exposure. Lung sulfide metabolite concentrations were transiently increased immediately after the end of the 3-h H2S exposure. Decreased cytochrome oxidase activity was observed in the olfactory epithelium following exposure to > or = 30 ppm H2S. Increased olfactory epithelial sulfide concentrations were observed following exposure to 400 ppm H2S. Hindbrain and nasal respiratory epithelial sulfide concentrations were unaffected by acute H2S exposure. Nasal respiratory epithelial cytochrome oxidase activity was reduced following acute exposure to > or = 30 ppm H2S. Liver sulfide concentrations were increased following exposure to > or = 200 ppm H2S and cytochrome oxidase activity was increased following inhalation exposure to > or = 10 ppm H2S. Our results suggest that cytochrome oxidase inhibition is a sensitive biomarker of H2S exposure in target tissues, and sulfide concentrations are unlikely to increase postexposure in the brain, lung, or nose following a single 3-h exposure to < or = 30 ppm H2S.     

Hydrogen sulfide suppresses the expression of MMP-8, MMP-13, and TIMP-1 in left ventricles of rats with cardiac volume overload

Aim:

To study the effects of hydrogen sulfide (H₂S) on the left ventricular expression of MMP-8, MMP-13, and TIMP-1 in a rat model of congenital heart disease.

Methods:

Male SD rats underwent abdominal aorta-inferior vena cava shunt operation. H₂S donor NaHS (56 μmol·kg⁻¹·d⁻¹, ip) was injected from the next day for 8 weeks. At 8 weeks, the hemodynamic parameters, including the left ventricular systolic pressure (LVSP), the left ventricular peak rate of contraction and relaxation (LV±dp/dt_max) and the left ventricular end diastolic pressure (LVEDP) were measured. The left ventricular tissues were dissected out, and hydroxyproline and collagen I contents were detected with ELISA. The expression of MMP-8, MMP-13, and a tissue inhibitor of metalloproteinase-1 (TIMP-1) in the tissues was measured using real-time PCR, Western blotting, and immunohistochemistry, respectively.

Results:

The shunt operation markedly reduced LVSP and LV±dp/dt_max, increased LVEDP, hydroxyproline and collagen I contents, as well as the mRNA and protein levels of MMP-8, MMP-13, and TIMP-1 in the left ventricles. Chronic treatment of the shunt operation rats with NaHS effectively prevented the abnormalities in the hemodynamic parameters, hydroxyproline and collagen I contents, and the mRNA and protein levels of MMP-13 and TIMP-1 in the left ventricles. NaHS also prevented the increase of MMP-8 protein expression, but did not affect the increase of mRNA level of MMP-8 in the shunt operation rats.

Conclusion:

H₂S suppresses protein and mRNA expression of MMP-8, MMP-13, and TIMP-1 in rats with cardiac volume overload, which may be contributed to the amelioration of ventricular structural remodeling and cardiac function.     

Dutch hospital drug formularies: pharmacotherapeutic variation and conservatism, but concurrence with national pharmacotherapeutic guidelines

AIMS: This research examines current hospital drug formularies (HDFs) of all Dutch general hospitals. It assesses the extent to which they recommend the same drugs, the breadth of their coverage in terms of therapeutic areas, drug groups incorporated and individuals drugs included, and their extent of conservatism by considering the year of introduction of the drugs included within groups. Furthermore, it considers the extent to which their recommendations concur and comply with those of national pharmacotherapeutic guidelines and the WHO Essential Drugs List (EDL). METHODS: Seventy-eight (81%) out of all 96 current Dutch HDFs were received of which 62 were suitable for study. Differences between HDFs and eventual associations with hospital characteristics were researched by statistical testing and case-control studies. To evaluate HDFs' concurrence with national guidelines and compliance with the WHO EDL, nine drug groups were studied in detail: benzodiazepines, calcium channel blockers, beta-adrenoceptor blocking agents, ACE-inhibitors, angiotensin-II inhibitors, NSAIDs, H2-receptor antagonists, 5HT3-antagonists, and H+-pump inhibitors. Concurrence and compliance with national guidelines and the WHO EDL was defined as inclusion of recommended drugs. Non-concurrence was defined as inclusion of nonrecommended drugs. RESULTS: The total number of indications addressed and drug groups incorporated within HDFs varied from 28 to 72 (median 56) and from 30 to 123 (median 97), respectively. The total number of individual drug entities (pharmacological substances) included ranged from 239 to 658 (median 430) and the total number of drug products, including all different dosage forms, from 412 to 1121 (median 655). Within drug groups, drug entities first marketed were most frequently included. Teaching hospitals were most likely to include recently marketed drugs. Depending on the drug group, HDFs' concurrence and compliance with national guidelines and the WHO EDL ranged from 35% to 100%. CONCLUSIONS: Findings indicate that Dutch HDFs are rather uniform in the indications addressed and the drug groups incorporated. However, the number of individual drug entities and drug products included within groups varies considerably. Furthermore, Dutch HDFs are considered rather conservative, as older drugs are favoured over more recent drugs. Generally, with some drug exceptions, Dutch HDFs concur and comply with recommendations in national pharmacotherapeutic guidelines and with the WHO EDL over 90%.