scholarly journals SPINK3 modulates mouse sperm physiology through the reduction of nitric oxide level independently of its trypsin inhibitory activity

Reproduction ◽  
2012 ◽  
Vol 143 (3) ◽  
pp. 281-295 ◽  
Author(s):  
L Zalazar ◽  
T E Saez Lancellotti ◽  
M Clementi ◽  
C Lombardo ◽  
L Lamattina ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Inhibitory Activity ◽  
Secretory Protein ◽  
No Donors ◽  
Mouse Sperm ◽  
Tyrosine Residues ◽  
Nitric Oxide Level ◽  
Exogenous Addition ◽  
Trypsin Inhibitory Activity

Serine protease inhibitor Kazal-type (SPINK3)/P12/PSTI-II is a small secretory protein from mouse seminal vesicle which contains a KAZAL domain and shows calcium (Ca2+)-transport inhibitory (caltrin) activity. This molecule was obtained as a recombinant protein and its effect on capacitated sperm cells was examined. SPINK3 inhibited trypsin activityin vitrowhile the fusion protein GST-SPINK3 had no effect on this enzyme activity. The inactive GST-SPINK3 significantly reduced the percentage of spermatozoa positively stained for nitric oxide (NO) with the specific probe DAF-FM DA and NO concentration measured by Griess method in capacitated mouse sperm; the same effect was observed when sperm were capacitated under low Ca2+concentration, using either intracellular (BAPTA-AM) or extracellular Ca2+(EDTA) chelators. The percentage of sperm showing spontaneous and progesterone-induced acrosomal reaction was significantly lower in the presence of GST-SPINK3 compared to untreated capacitated spermatozoa. Interestingly, this decrease was overcome by the exogenous addition of the NO donors, sodium nitroprusside (SNP), andS-nitrosoglutathione (GSNO). Phosphorylation of sperm proteins in tyrosine residues was partially affected by GST-SPINK3, however, only GSNO was able to reverse this effect. Sperm progressive motility was not significantly diminished by GST-SPINK3 or BAPTA-AM but enhanced by the addition of SNP. This is the first report that demonstrates that SPINK3 modulates sperm physiology through a downstream reduction of endogenous NO concentration and independently of SPINK3 trypsin inhibitory activity.

scholarly journals In vitro anticancer activity of hydrogen sulfide and nitric oxide alongside nickel nanoparticle and novel mutations in their genes in CRC patients

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Zjwan Housein ◽  
Tayeb Sabir Kareem ◽  
Abbas Salihi
Keyword(s):  
Nitric Oxide ◽  
Cell Proliferation ◽  
Hydrogen Sulfide ◽  
Direct Sequencing ◽  
Scorpion Venom ◽  
Enos Gene ◽  
Cytotoxic Activities ◽  
No Donors ◽  
The Impact

AbstractThis study was carried out to assess the impact of nickel nanoparticles (NiNPs) as well as scorpion venom on colorectal cancer (CRC) cells in the presence and/or absence of 5-fluorouracil (5-FU), hydrogen sulfide (H2S), and nitric oxide (NO) donors and to determine alterations in endothelial NO synthase (eNOS) and cystathionine γ-lyase (CSE) enzyme-producing genes in CRC patients. The IC50 of both H2S and NO donors, along with NiNPs, were determined. The CRC cells were treated for 24hrs, and the cytotoxic activities were assessed using the MTT test. Moreover, the apoptosis was determined after 24hrs and 48hrs using TUNEL assay. Furthermore, the mutations in the eNOS gene (intron 4, -786T>C and 894 G>T) and CSE gene (1364GT) were determined using direct sequencing. The IC50 values for sodium disulfide (Na2S) and sodium nitroprusside (SNP) at 24hrs treatment were found to be 5 mM and 10−6 M, respectively, while the IC50 value for 5-FU was reached after 5-days of treatment in CRC cell line. Both black and yellow scorpion venoms showed no inhibition of cell proliferation after 24hrs treatment. Furthermore, Na2S showed a significant decrease in cell proliferation and an increase in apoptosis. Moreover, a co-treatment of SNP and 5-FU resulted in inhibition of the cytotoxic effect of 5-FU, while a combination treatment of NiNPs with Na2S, SNP, and 5-FU caused highly significant cytotoxicity. Direct sequencing reveals new mutations, mainly intronic variation in eNOS gene that has not previously been described in the database. These findings indicate that H2S promotes the anticancer efficiency of 5-FU in the presence of NiNPs while NO has antiapoptotic activity in CRC cell lines.

Diarylheptanoids with In Vitro Inducible Nitric Oxide Synthesis Inhibitory Activity from Alnus hirsuta

Planta Medica ◽  
2000 ◽  
Vol 66 (6) ◽  
pp. 551-553 ◽  
Author(s):  
Min-Won Lee ◽  
Na-Young Kim ◽  
Myung-Shin Park ◽  
Kyoung-Hwan Ahn ◽  
Sang-Hak Toh ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Inhibitory Activity ◽  
Nitric Oxide Synthesis ◽  
Alnus Hirsuta ◽  
Inducible Nitric Oxide

Evidence that nitric oxide synthase is involved in progesterone-induced acrosomal exocytosis in mouse spermatozoa

1997 ◽  
Vol 9 (4) ◽  
pp. 433 ◽  
Author(s):  
María Beléen Herrero ◽  
J. Marcelo Viggiano ◽  
Silvina Pérez Martínez ◽  
Martha F. de Gimeno
Keyword(s):  
Nitric Oxide ◽  
Methyl Ester ◽  
Nitric Oxide Synthase ◽  
No Synthase ◽  
Nitric Oxide Donor ◽  
Mouse Sperm ◽  
Acrosomal Exocytosis ◽  
Oxide Synthase ◽  
Spermine Nonoate

In a recent work, we detected nitric oxide synthase (NO synthase) in the acrosome and tail of mouse and human spermatozoa by an immunofluorescence technique. Also, NO-synthase inhibitors added during sperm capacitationin vitro reduced the percentage of oocytes fertilized in vitro, suggesting a role for NO synthase in sperm function. Therefore, in the present study the effect of three NO-synthase inhibitors, NG-nitro-L-arginine methyl ester (L-NAME), NG-nitro-D-arginine methyl ester (D-NAME) and L-NG-nitro-arginine (NO2-arg), and of a nitric oxide donor, spermine-NONOate, on the progesterone-induced acrosome reaction of mouse sperm was examined. NO-synthase inhibitors were added at 0, 60 or 90 min during capacitation; at 120 min, mouse epididymal spermatozoa were exposed to 15 µM progesterone for another 15 min. In another set of experiments, different concentrations of spermine-NONOate were added to capacitated spermatozoa for 15 min; in these experiments, progesterone was not included. NO2-arg and L-NAME blocked progesterone-induced exocytosis regardless of the time at which these inhibitors were added. Moreover, D-NAME did not inhibit exocytosis. In contrast, spermine-NONOate stimulated the acrosomal exocytosis in vitro directly. These results provide evidence that mouse sperm NO synthase participates in the progesterone-induced acrosome reactionin vitro and that nitric oxide induces this event.

Restoring Oxygen Carrying Capacity of Sickle Erythrocytes with Nitric Oxide Donors.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3785-3785
Author(s):  
Borys Hrinczenko
Keyword(s):  
Nitric Oxide ◽  
Oxygen Affinity ◽  
Red Cell ◽  
No Donors ◽  
Sickle Erythrocytes ◽  
Low Levels ◽  
Pre Treatment ◽  
Normal Controls

Abstract Sickle cell anemia (SCA) is an inherited blood disorder of hemoglobin function. A genetic mutation results in the substitution of a valine for glutamic acid residue at position 6 of the beta-globin chain yielding the mutant hemoglobin S (HbS). HbS polymerizes within erythrocytes during deoxygenation resulting in altered affinity of oxygen binding. The slightly different P50 (PO2 at which Hb is half-saturated with oxygen) values of sickle erythrocytes obtained during either oxygenation or deoxygenation (hysteresis) demonstrate HbS polymerization induced inhibition of oxygen affinity. Nitric oxide (NO) has been found to be an important signaling molecule in the circulatory system. NO derivatives of Hb provide insights into the physiological role of Hb. NO can bind to Hb at either the heme moiety forming nitrosylhemoglobin (HbNO) or to the conserved beta-93 cysteine yielding S-nitrosohemoglobin (SNO-Hb). In deoxygenated venous blood NO preferentially binds to the hemes of Hb forming HbNO while in oxygenated arterial blood NO binds to the beta-93 cysteine residues forming SNO-Hb. Increased oxygen affinity is seen in both SNO-Hb (Bonaventura C, et al, 1999) and also with HbNO. Decreasing the HbS P50 inhibits intra-erythrocyte HbS polymerization that may be an effective strategy to treat SCA. Clinical trials of NO breathing effects on oxygen affinity are conflicting. One study found an increased oxygen affinity of blood from SCA patients breathing 80 ppm NO with no effect seen in normal controls (Head A, et al, 1997). Another study found that levels of NO bound to Hb are too low to affect overall oxygen affinity (Gladwin M, et al, 1999). The purpose of this in vitro study was to determine the oxygen affinity of deoxygenated sickle erythrocytes pre-treated with exogenous NO donors. Blood from SCA (HbSS) and normal controls (HbAA) were collected and suspended in PBS buffer and deoxygenated with argon gas. The Hb concentration of each sample was calculated and then was either left untreated (control) or treated with varying concentrations of NO donors. The NO donors included: 2-(N, N-diethylamino)-diazenolate-2-oxide (DEANO), S-nitroso-N-acetylpenicillamine (SNAP), sodium nitroprusside (SNP), an aqueous solution of NO, and sodium trioxodinitrate (Angeli’s salt, AS). Methemoglobin and protein degradation were negligible. Samples were then transferred via airtight syringes into a stirred and temperature controlled (37°C) chamber of PBS solution at ambient oxygen pressure fitted with a very sensitive oxygen electrode. Oxygen levels were measured in real time. The amount of oxygen extracted from the PBS medium followed first order kinetics. Studies with HbSS red cell suspensions showed that the largest increment in oxygen extraction from the medium was obtained with DEANO pre-treatment. Calculations indicated that low levels of NO treatment, at approximately a 1:1000 ratio of [NO]/[heme], yielded the largest oxygen consumption. The effects of pre-treatment with the other NO donors on sickle erythrocytes (HbSS) were not as pronounced. DEANO is an NO donor yielding a “pure” NO radical as opposed to other redox forms. Similar studies with HbAA and HbSC did not show increases in oxygen extraction. Taken together the data suggest that low levels of NO perturb the quaternary structure of intraerythrocyte HbS polymer allowing depolymerization and increased oxygen affinity. The hope is that these in vitro studies will better characterize the role of NO in its interactions with Hb and the red cell and to use this knowledge for potential therapies in diseases such as SCA.

In Vitro Activation of Soluble Guanylyl Cyclase and Nitric Oxide Release:  A Comparison of NO Donors and NO Mimetics†

Biochemistry ◽  
2001 ◽  
Vol 40 (31) ◽  
pp. 9256-9264 ◽  
Author(s):  
Jennifer D. Artz ◽  
Violeta Toader ◽  
Sergei I. Zavorin ◽  
Brian M. Bennett ◽  
Gregory R. J. Thatcher
Keyword(s):  
Nitric Oxide ◽  
Guanylyl Cyclase ◽  
Soluble Guanylyl Cyclase ◽  
No Donors ◽  
Nitric Oxide Release ◽  
In Vitro Activation

scholarly journals Involvement of nitric oxide donor compounds in the bactericidal activity of human neutrophils in vitro

2003 ◽  
Vol 52 (4) ◽  
pp. 303-308 ◽  
Author(s):  
Magdalena Klink ◽  
Maciej Cedzyński ◽  
Anna St Świerzko ◽  
Henryk Tchórzewski ◽  
Zofia Sulowska
Keyword(s):  
Nitric Oxide ◽  
Coronary Heart Disease ◽  
Heart Disease ◽  
Bactericidal Activity ◽  
Nitric Oxide Donor ◽  
Human Neutrophils ◽  
Bacterial Strains ◽  
No Donor ◽  
No Donors

The bactericidal activity of human neutrophils against extracellular and facultatively intracellular bacteria was studied in the presence of the nitric oxide (NO) donors sodium nitroprusside (SNP) and 3-morpholinosydnonimine (SIN-1), a molsidomine metabolite. SNP and molsidomine are drugs commonly used as nitrovasodilators in coronary heart disease. It is demonstrated here that the NO donor compounds themselves did not affect the viability and survival of the bacterial strains tested. Neither SNP nor SIN-1 had any effect on the process of bacteria ingestion. In contrast, NO donors enhanced the ability of neutrophils to kill Escherichia coli, Proteus vulgaris and Salmonella Anatum. However, strains differed in their susceptibility to SNP- and SIN-1-mediated killing by neutrophils. Removal of the superoxide anion reduced the bactericidal activity of SNP- and SIN-1-treated neutrophils against E. coli and S. Anatum. This suggests that the NO derivatives formed in the reaction of NO generated from donors with the reactive oxygen species released by phagocytosed neutrophils potentiate the bactericidal activity of human neutrophils in vitro. The above original observation discussed here suggests clinical significance for the treatment of patients with nitrovasodilators in the course of coronary heart disease therapy.

NITROSYL FERREDOXINES MIMETICS AS PHOSPHODIESTERASE INHIBITORS: SYNTHESIS, STRUCTURE AND ACTIVITY IN VITRO

2021 ◽  
Author(s):  
Natalia Sanina ◽  
Keyword(s):  
Nitric Oxide ◽  
Solid Phase ◽  
Phosphodiesterase Inhibitors ◽  
Low Molecular Weight ◽  
Structure And Properties ◽  
No Donors ◽  
Low Molecular Weight Iron ◽  
Dinitrosyl Complexes ◽  
Structure And Activity

The paper presents and discusses experimental data on the synthesis, study of the structure and properties in the solid phase and in solutions of new low-molecular-weight iron dinitrosyl complexes with N,S-ligands of the 1,2,4 -1-H-triazole N,S-ligands of the“g = 2.03” family, nitric oxide (NO) donors, and effective inhibitors of cGMP phosphodiesterases.

scholarly journals Inducible nitric oxide synthase (NOS2) expressed in septic patients is nitrated on selected tyrosine residues: implications for enzymic activity

2002 ◽  
Vol 366 (2) ◽  
pp. 399-404 ◽  
Author(s):  
Sophie LANONE ◽  
Philippe MANIVET ◽  
Jacques CALLEBERT ◽  
Jean-Marie LAUNAY ◽  
Didier PAYEN ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Inducible Nitric Oxide Synthase ◽  
Protein Modification ◽  
Enzymic Activity ◽  
Tyrosine Residues ◽  
Oxide Synthase ◽  
Inducible Nitric Oxide

Tyrosine nitration is a post-translational protein modification with potentially significant biological implications. In the present study we demonstrate, for the first time, that tyrosine residues of human inducible nitric oxide synthase (NOS2) can be nitrated by peroxynitrite in vitro, leading to a decreased activity. Moreover, we show that NOS2 expressed in a skeletal muscle from septic patients is nitrated on selective tyrosine residues belonging to a canonic sequence. This phenomenon could be an endogenous mechanism of in vivo modulation of NOS2 enzymic activity.

scholarly journals Effects of nanosuspension and inclusion complex techniques on the in vitro protease inhibitory activity of naproxen

2014 ◽  
Vol 50 (1) ◽  
pp. 165-171 ◽  
Author(s):  
Senthil Rajan Dharmalingam ◽  
Kumarappan Chidambaram ◽  
Srinivasan Ramamurthy ◽  
Shamala Nadaraju
Keyword(s):  
Inclusion Complex ◽  
Inhibitory Activity ◽  
Lower Percentage ◽  
Physiochemical Properties ◽  
Trypsin Inhibitory Activity ◽  
Inflammatory Drugs ◽  
Anti Inflammatory ◽  
Antitryptic Activity ◽  
Inflammatory Effect

This study investigated the effects of nanosuspension and inclusion complex techniques on in vitro trypsin inhibitory activity of naproxen—a member of the propionic acid derivatives, which are a group of antipyretic, analgesic, and non-steroidal anti-inflammatory drugs. Nanosuspension and inclusion complex techniques were used to increase the solubility and anti-inflammatory efficacy of naproxen. The evaporative precipitation into aqueous solution (EPAS) technique and the kneading methods were used to prepare the nanosuspension and inclusion complex of naproxen, respectively. We also used an in vitro protease inhibitory assay to investigate the anti-inflammatory effect of modified naproxen formulations. Physiochemical properties of modified naproxen formulations were analyzed using UV, IR spectra, and solubility studies. Beta-cyclodextrin inclusion complex of naproxen was found to have a lower percentage of antitryptic activity than a pure nanosuspension of naproxen did. In conclusion, nanosuspension of naproxen has a greater anti-inflammatory effect than the other two tested formulations. This is because the nanosuspension formulation reduces the particle size of naproxen. Based on these results, the antitryptic activity of naproxen nanosuspension was noteworthy; therefore, this formulation can be used for the management of inflammatory disorders.

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