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PubMed1998

Increased Cat3-mediated cationic amino acid transport functionally compensates in Cat1 knockout cell lines.

Arginine transport is important for a number of biological processes in vertebrates, and its transport may be rate-limiting for the production of nitric oxide. The majority of L-Arg transport is mediated by System y+, although several other carriers have been kinetically defined. System y+ cationic amino acid transport is mediated by proteins encoded by a family of genes, Cat1, Cat2, and Cat3. High affinity L-arginine transport was investigated in embryonic fibroblast cells derived from Cat1 knockout mice that lack functional Cat1. Both wild type and knockout cells transport arginine with comparable Km and Vmax. However, the apparent affinity for lysine transport was 2.4 times lower in Cat1(-/-) cells when compared with wild type cells, a property characteristic of Cat3-mediated transport. Northern analysis-documented Cat2 mRNA increased 2-fold, whereas Cat3 mRNA levels increased 11-fold in Cat1(-/-) relative to Cat1(+/+) cells. The low affinity Cat2a mRNA was not detectably expressed in these cells. Even though Cat3 expression is normally limited to adult brain, there was a large increase in the amount of Cat3 protein present at the plasma membrane of Cat1(-/-) embryonic fibroblast cells. These results suggest that Cat3 compensates for the loss of functional Cat1 in cells derived from Cat1 knockout mice and mediates the majority of high affinity arginine transport.

Amino Acid Transport Systems, Basic↗

PubMed1998

Loss of antinociception induced by naloxone benzoylhydrazone in nociceptin receptor-knockout mice.

Nociceptin and nociceptin receptor, which show structural similarities to opioid peptides and opioid receptors, respectively, have been recently found to constitute a novel neuromodulatory system. In the brain, however, the physiological role of the modulation via the nociceptin receptor is still unclear. Administered nociceptin produces hyperalgesia and hypolocomotion, whereas the nociceptin receptor-knockout mice show no significant abnormalities in nociceptive thresholds and locomotion. To clarify possible involvement of the nociceptin receptor in the regulation of nociception and locomotion, we made use of the knockout mice and naloxone benzoylhydrazone (NalBzoH) identified originally as a ligand for opioid receptors. Experiments on the cultured cells transfected with the nociceptin receptor cDNA showed that NalBzoH competed with [3H]nociceptin binding and attenuated the nociceptin-induced inhibition of cAMP accumulation. Furthermore, behavioral studies demonstrated that NalBzoH completely inhibited nociceptin-induced hyperalgesia and hypolocomotion. It is therefore likely that NalBzoH can act as a potent antagonist for the nociceptin receptor in vivo. In wild-type mice, NalBzoH induced antinociception but did not affect locomotor activity. In contrast, in the knockout mice, no significant changes in nociception and locomotion were induced by NalBzoH. These results clearly suggest that the nociceptin system takes part in the physiological regulation of nociceptive thresholds but not in the basal modulation of locomotion.

Animals↗

PubMed1999

Liver-specific inactivation of the abetalipoproteinemia gene completely abrogates very low density lipoprotein/low density lipoprotein production in a viable conditional knockout mouse.

Conventional knockout of the microsomal triglyceride transfer protein large subunit (lMTP) gene is embryonic lethal in the homozygous state in mice. We have produced a conditional lMTP knockout mouse by inserting loxP sequences flanking exons 5 and 6 by gene targeting. Homozygous floxed mice were born live with normal plasma lipids. Intravenous injection of an adenovirus harboring Cre recombinase (AdCre1) produced deletion of exons 5 and 6 and disappearance of lMTP mRNA and immunoreactive protein in a liver-specific manner. There was also disappearance of plasma apolipoprotein (apo) B-100 and marked reduction in apoB-48 levels. Wild-type mice showed no response, and heterozygous mice, an intermediate response, to AdCre1. Wild-type mice doubled their plasma cholesterol level following a high cholesterol diet. This hypercholesterolemia was abolished in AdCre1-treated lMTP-/- mice, the result of a complete absence of very low/intermediate/low density lipoproteins and a slight reduction in high density lipoprotein. Heterozygous mice showed an intermediate lipoprotein phenotype. The rate of accumulation of plasma triglyceride following Triton WR1339 treatment in lMTP-/- mice was <10% that in wild-type animals, indicating a failure of triglyceride-rich lipoprotein production. Pulse-chase experiments using hepatocytes isolated from wild-type and lMTP-/- mice revealed a failure of apoB secretion in lMTP-/- animals. Therefore, the liver-specific inactivation of the lMTP gene completely abrogates apoB-100 and very low/intermediate/low density lipoprotein production. These conditional knockout mice are a useful in vivo model for studying the role of MTP in apoB biosynthesis and the biogenesis of apoB-containing lipoproteins.

Abetalipoproteinemia↗

PubMed2006

Ocular inflammatory responses in the EP2 and EP4 receptor knockout mice.

PURPOSE: To examine the role of EP2 and EP4 receptors in murine ocular inflammation. METHODS: Prostaglandin EP2 and EP4 receptor knockout and wild-type mice were treated topically with prostaglandin E2, SDF-1, and RANTES and lipopolysaccharide by intravitreal injection. Paracentesis was performed by puncturing the cornea. The increase in the level of aqueous humor protein and the number of leukocytes were measured and the vascular leakage of protein was visualized using fluorescein angiography. RESULTS: In the EP2 receptor knockout mice, there was significant inhibition of the disruption of the blood-aqueous barrier caused by lipopolysaccharides, paracentesis, prostaglandin E2, SDF-1, and RANTES. Reductions in the disruption in the blood-aqueous barrier and leukocyte infiltration after lipopolysaccharide injection and paracentesis were significant, but there was no increase in the aqueous humor protein level after prostaglandin E2 treatment in EP4 receptor knockout mice. CONCLUSIONS: The results of the present experiments suggest that EP2 and EP4 receptors partly mediate the disruption of the blood-aqueous barrier and leukocyte infiltration induced by prostaglandin E2, SDF-1, RANTES, and lipopolysaccharides.

Animals↗

PubMed2001

Residual Cajal bodies in coilin knockout mice fail to recruit Sm snRNPs and SMN, the spinal muscular atrophy gene product.

Cajal bodies (CBs) are nuclear suborganelles involved in the biogenesis of small nuclear ribonucleoproteins (snRNPs). In addition to snRNPs, they are highly enriched in basal transcription and cell cycle factors, the nucleolar proteins fibrillarin (Fb) and Nopp140 (Nopp), the survival motor neuron (SMN) protein complex, and the CB marker protein, p80 coilin. We report the generation of knockout mice lacking the COOH-terminal 487 amino acids of coilin. Northern and Western blot analyses demonstrate that we have successfully removed the full-length coilin protein from the knockout animals. Some homozygous mutant animals are viable, but their numbers are reduced significantly when crossed to inbred backgrounds. Analysis of tissues and cell lines from mutant animals reveals the presence of extranucleolar foci that contain Fb and Nopp but not other typical nucleolar markers. These so-called "residual" CBs neither condense Sm proteins nor recruit members of the SMN protein complex. Transient expression of wild-type mouse coilin in knockout cells results in formation of CBs and restores these missing epitopes. Our data demonstrate that full-length coilin is essential for proper formation and/or maintenance of CBs and that recruitment of snRNP and SMN complex proteins to these nuclear subdomains requires sequences within the coilin COOH terminus.

Animals↗

PubMed2000

Impaired preneoplastic changes and liver tumor formation in tumor necrosis factor receptor type 1 knockout mice.

Hepatic stem cells (oval cells) proliferate within the liver after exposure to a variety of hepatic carcinogens and can generate both hepatocytes and bile duct cells. Oval cell proliferation is commonly seen in the preneoplastic stages of liver carcinogenesis, often accompanied by an inflammatory response. Tumor necrosis factor (TNF), an inflammatory cytokine, is also important in liver regeneration and hepatocellular growth. The experiments reported here explore the relationship among the TNF inflammatory pathway, liver stem cell activation, and tumorigenesis. We demonstrate that TNF is upregulated during oval cell proliferation induced by a choline-deficient, ethionine-supplemented diet and that it is expressed by oval cells. In TNF receptor type 1 knockout mice, oval cell proliferation is substantially impaired and tumorigenesis is reduced. Oval cell proliferation is impaired to a lesser extent in interleukin 6 knockout mice and is unchanged in TNF receptor type 2 knockout mice. These findings demonstrate that TNF signaling participates in the proliferation of oval cells during the preneoplastic phase of liver carcinogenesis and that loss of signaling through the TNF receptor type 1 reduces the incidence of tumor formation. The TNF inflammatory pathway may be a target for therapeutic intervention during the early stages of liver carcinogenesis.

Animals↗

PubMed2001

Gap junctional coupling in lenses from alpha(8) connexin knockout mice.

Lens fiber cell gap junctions contain alpha(3) (Cx46) and alpha(8) (Cx50) connexins. To examine the roles of the two different connexins in lens physiology, we have genetically engineered mice lacking either alpha(3) or alpha(8) connexin. Intracellular impedance studies of these lenses were used to measure junctional conductance and its sensitivity to intracellular pH. In Gong et al. 1998, we described results from alpha(3) connexin knockout lenses. Here, we present original data from alpha(8) connexin knockout lenses and a comparison with the previous results. The lens has two functionally distinct domains of fiber cell coupling. In wild-type mouse lenses, the outer shell of differentiating fibers (see 1, DF) has an average coupling conductance per area of cell-cell contact of approximately 1 S/cm(2), which falls to near zero when the cytoplasm is acidified. In the inner core of mature fibers (see 1, MF), the average coupling conductance is approximately 0.4 S/cm(2), and is insensitive to acidification of the cytoplasm. Both connexin isoforms appear to contribute about equally in the DF since the coupling conductance for either heterozygous knockout (+/-) was approximately 70% of normal and 30-40% of the normal for both -/- lenses. However, their contribution to the MF was different. About 50% of the normal coupling conductance was found in the MF of alpha(3) +/- lenses. In contrast, the coupling of MF in the alpha(8) +/- lenses was the same as normal. Moreover, no coupling was detected in the MF of alpha(3) -/- lenses. Together, these results suggest that alpha(3) connexin alone is responsible for coupling MF. The pH- sensitive gating of DF junctions was about the same in wild-type and alpha(3) connexin -/- lenses. However, in alpha(8) -/- lenses, the pure alpha(3) connexin junctions did not gate closed in the response to acidification. Since alpha(3) connexin contributes about half the coupling conductance in DF of wild-type lenses, and that conductance goes to zero when the cytoplasmic pH drops, it appears alpha(8) connexin regulates the gating of alpha(3) connexin. Both connexins are clearly important to lens physiology as lenses null for either connexin lose transparency. Gap junctions in the MF survive for the lifetime of the organism without protein turnover. It appears that alpha(3) connexin provides the long-term communication in MF. Gap junctions in DF may be physiologically regulated since they are capable of gating when the cytoplasm is acidified. It appears alpha(8) connexin is required for gating in DF.

Animals↗

PubMed2005

Thrombopoietin production in wild-type and interleukin-6 knockout mice with acute inflammation.

Clinical and laboratory studies indicate that thrombopoietin (TPO) gene expression increases during inflammation. To clarify the role of interleukin 6 (IL-6) in this process, blood cell counts, plasma TPO concentrations, and hepatic and renal TPO mRNA levels were investigated in wild-type and IL-6 knockout mice, with sterile abscesses produced by subcutaneous injection of turpentine oil. Treatment did not cause a change in blood cell counts during the 72 h period of observation. The numbers of thrombocytes and erythrocytes were slightly lower in the IL-6 knockout mice than in the wild-type littermates under all conditions. Plasma IL-6 and TPO concentrations increased on turpentine injection only in the wild-type mice. In addition, turpentine treatment of these caused an increase in hepatic TPO mRNA levels as assessed by competitive polymerase chain reaction (RT-PCR) and real-time PCR, whereas renal TPO mRNA levels were unaltered. TPO mRNA levels did not increase in the livers of IL-6 knockout mice on turpentine treatment. These results support the concept that TPO behaves like an acute-phase protein in that its synthesis is induced by IL-6 in the liver.

Acute Disease↗

PubMed2000

Heterozygous DNA mismatch repair gene PMS2-knockout mice are susceptible to intestinal tumor induction with N-methyl-N-nitrosourea.

PMS2-deficient (PMS2(-/-)) mice are hypersensitive to N-methyl-N-nitrosourea (MNU)-induced thymic lymphomas based on the failure to initiate mismatch repair (MMR) at O(6)-methylguanine:T mismatches formed after MNU exposure. However, heterozygous PMS2 knockout (PMS2(+/-)) mice do not develop spontaneous tumors, suggesting that they have sufficient MMR function to prevent genomic instability. We hypothesized that in PMS2(+/-) mice, exogenous carcinogens may either mutationally knockout the remaining normal allele leading cells to develop tumors or introduce sufficient DNA adducts and mismatches to overload the lower capacity for MMR, leading in either case to an increased rate of tumor production. In the present study, PMS2(+/-) mice and their littermate PMS2(+/+) mice were monitored for tumor incidence following MNU treatment. Mice were given 50 mg MNU/kg i.p. when 5 weeks old. They demonstrated a similar incidence of thymic lymphomas, suggesting that expression of the single normal PMS2 allele is sufficient to protect the thymus and implying that a single dose of MNU may not efficiently knock out the remaining PMS2 allele in the thymus. Surprisingly, PMS2(+/-) mice were significantly more likely to develop intestinal tumors-both adenomas and adenocarcinomas-after MNU than were PMS2(+/+) mice (2.34 +/- 0.34 tumors per mouse versus 1.34 +/- 0.25 tumors per mouse; P < 0.05). The intestinal tumors were located mainly in the small intestine. However, these tumors in both the PMS2(+/-) mice and PMS2(+/+) mice did not show microsatellite instability characteristic of loss of MMR. These results suggest that a single normal PMS2 allele can protect thymus but not intestine from MNU carcinogenesis. Organ-specific factors might influence MMR- mediated resistance to methylating agents. Heterozygous PMS2 knockout mice may be used as a promising animal model for intestinal tumorigenesis studies involving environmental carcinogens.

Adenosine Triphosphatases↗

PubMed2001

BACE knockout mice are healthy despite lacking the primary beta-secretase activity in brain: implications for Alzheimer's disease therapeutics.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by accumulation of amyloid plaques and neurofibrillary tangles in the brain. The major components of plaque, beta-amyloid peptides (Abetas), are produced from amyloid precursor protein (APP) by the activity of beta- and gamma-secretases. beta-secretase activity cleaves APP to define the N-terminus of the Abeta1-x peptides and, therefore, has been a long- sought therapeutic target for treatment of AD. The gene encoding a beta-secretase for beta-site APP cleaving enzyme (BACE) was identified recently. However, it was not known whether BACE was the primary beta-secretase in mammalian brain nor whether inhibition of beta-secretase might have effects in mammals that would preclude its utility as a therapeutic target. In the work described herein, we generated two lines of BACE knockout mice and characterized them for pathology, beta-secretase activity and Abeta production. These mice appeared to develop normally and showed no consistent phenotypic differences from their wild-type littermates, including overall normal tissue morphology and brain histochemistry, normal blood and urine chemistries, normal blood-cell composition, and no overt behavioral and neuromuscular effects. Brain and primary cortical cultures from BACE knockout mice showed no detectable beta-secretase activity, and primary cortical cultures from BACE knockout mice produced much less Abeta from APP. The findings that BACE is the primary beta-secretase activity in brain and that loss of beta-secretase activity produces no profound phenotypic defects with a concomitant reduction in beta-amyloid peptide clearly indicate that BACE is an excellent therapeutic target for treatment of AD.

Alzheimer Disease↗

PubMed1998

Generalized glycogen storage and cardiomegaly in a knockout mouse model of Pompe disease.

Glycogen storage disease type II (GSDII; Pompe disease), caused by inherited deficiency of acid alpha-glucosidase, is a lysosomal disorder affecting heart and skeletal muscles. A mouse model of this disease was obtained by targeted disruption of the murine acid alpha-glucosidase gene (Gaa) in embryonic stem cells. Homozygous knockout mice (Gaa -/-) lack Gaa mRNA and have a virtually complete acid alpha-glucosidase deficiency. Glycogen-containing lysosomes are detected soon after birth in liver, heart and skeletal muscle cells. By 13 weeks of age, large focal deposits of glycogen have formed. Vacuolar spaces stain positive for acid phosphatase as a sign of lysosomal pathology. Both male and female knockout mice are fertile and can be intercrossed to produce progeny. The first born knockout mice are at present 9 months old. Overt clinical symptoms are still absent, but the heart is typically enlarged and the electrocardiogram is abnormal. The mouse model will help greatly to understand the pathogenic mechanism of GSDII and is a valuable instrument to explore the efficacy of different therapeutic interventions.

Animals↗

PubMed2006

KCNJ11 gene knockout of the Kir6.2 KATP channel causes maladaptive remodeling and heart failure in hypertension.

Heart failure is a growing epidemic, with systemic hypertension a major risk factor for development of disease. However, the molecular determinants that prevent the transition from a state of hypertensive load to that of overt cardiac failure remain largely unknown. Here in experimental hypertension, knockout of the KCNJ11 gene, encoding the Kir6.2 pore-forming subunit of the sarcolemmal ATP-sensitive potassium (K(ATP)) channel, predisposed to heart failure and death. Defective decoding of hypertension-induced metabolic distress signals in the K(ATP) channel knockout set in motion pathological calcium overload and aggravated cardiac remodeling through a calcium/calcineurin-dependent cyclosporine-sensitive pathway. Rescue of the failing K(ATP) knockout phenotype was achieved by alternative control of myocardial calcium influx, bypassing uncoupled metabolic-electrical integration. The intact KCNJ11-encoded K(ATP) channel is thus a required safety element preventing hypertension-induced heart failure, with channel dysfunction a molecular substrate for stress-associated channelopathy in cardiovascular disease.

Animals↗

PubMed2006

Polymeric IgR knockout mice are more susceptible to mycobacterial infections in the respiratory tract than wild-type mice.

It is generally accepted that cellular, and not humoral immunity, plays the crucial role in defense against intracellular bacteria. However, accumulating data indicate the importance of humoral immunity for the defense against a number of intracellular bacteria, including mycobacteria. We have investigated the role of secretory IgA, the main isotype found in mucosal tissues, in protection against mycobacterial infection, using polymeric IgR (pIgR)-deficient mice. Characterization of the humoral response induced after intra-nasal immunizations with the mycobacterial antigen PstS-1 revealed a loss of antigen-specific IgA response in saliva from the knockout mice. IgA level in the bronchoalveolar lavage of knockout mice was similar to wild-type level, although the IgA antibodies must have reached the lumen by other means than pIgR-mediated transport. Infection with Mycobacterium bovis bacillus Calmette-Guérin (BCG) demonstrated that the immunized pIgR-/- mice were more susceptible to BCG infection than immunized wild-type mice, based on higher bacterial loads in the lungs. This was accompanied by a reduced production of both IFN-gamma and tumor necrosis factor-alpha (TNF-alpha) in the lungs. Additionally, the pIgR-/- mice displayed reduced natural resistance to mycobacterial infection proved by significantly higher bacterial growth in their lungs compared with wild-type mice after infection with virulent Mycobacterium tuberculosis. The knockout mice appeared to have a delayed mycobacteria-induced immune response with reduced expression of protective mediators, such as IFN-gamma, TNF-alpha, inducible nitric oxide synthase and regulated upon activation normal T cell sequence, during early infection. Collectively, our results show that actively secreted IgA plays a role in protection against mycobacterial infections in the respiratory tract, by blocking entrance of bacilli into the lungs, in addition to modulation of the mycobacteria-induced pro-inflammatory response.

Administration, Intranasal↗

PubMed1999

A system for rapid generation of coat color-tagged knockouts and defined chromosomal rearrangements in mice.

Gene targeting in mouse embryonic stem (ES) cells can be used to generate single gene mutations or defined multi-megabase chromosomal rearrangements when applied with the Cre- loxP recombination system. While single knockouts are essential for uncovering functions of cloned genes, chromosomal rearrangements are great genetic tools for mapping, mutagenesis screens and functional genomics. The conventional approach to generate mice with targeted alterations of the genome requires extensive molecular cloning to build targeting vectors and DNA-based genotyping for stock maintenance. Here we describe the design and construction of a two-library system to facilitate high throughput gene targeting and chromo-somal engineering. The unique feature of these libraries is that once a clone is isolated, it is essentially ready to be used for insertional targeting in ES cells. The two libraries each bear a complementary set of genetic markers tailored so that the vector can be used for Cre- loxP -based chromosome engineering as well as single knockouts. By incorporating mouse coat color markers into the vectors, we illustrate a widely applicable method for stock maintenance of ES cell-derived mice with single gene knockouts or more extensive chromosomal rearrangements.

Agouti Signaling Protein↗

PubMed1996

Application of primary hepatocytes from p53-knockout mice for studies of expression of Cyp3a.

CYP3A rapidly disappears in primary hepatocytes, although the primary cells are suitable for studies of the regulation of CYP3A genes. In the present study, we found that Cyp3a mRNA could be expressed in the primary hepatocytes from p53-knockout mice for at least 2 weeks when the cells were cultured in the presence of dexamethasone. Propoxycoumarin O-depropylase activity, which is known to be mainly catalyzed by CYP3A, was maintained at a level of 50% of the initial activity even after 5 days of culture, and the activity correlated with the expression level of Cyp3a mRNA in the primary hepatocytes from p53-knockout mice. The cells remained morphologically intact during 4 weeks. These results suggest that hepatocytes from p53-knockout mice are a useful tool for studies of the expression of Cyp3a.

7-Alkoxycoumarin O-Dealkylase↗

PubMed2001

High sensitivity of Nrf2 knockout mice to acetaminophen hepatotoxicity associated with decreased expression of ARE-regulated drug metabolizing enzymes and antioxidant genes.

Nrf2, which belongs to the basic leucine zipper (bZip) transcription factor family, has been implicated as a key molecule involved in antioxidant-responsive element (ARE)-mediated gene expression. In order to examine the role of Nrf2 in protection against xenobiotic toxicity, the sensitivity of nrf2 knockout mice to acetaminophen (N-acetyl-4-aminophenol (APAP)) was analyzed. The saturation of detoxification pathways after high levels of exposure to APAP is known to induce hepatotoxicity. Two factors important in its detoxification are UDP-glucuronosyltransferase (UDP-GT), an ARE-regulated phase-II drug-metabolizing enzyme, and glutathione (GSH), an antioxidant molecule whose synthesis depends on ARE-regulated gamma-glutamylcysteine synthetase (gammaGCS). Two- to 4-month-old male mice were orally administered a single dose of APAP at 0, 150, 300, or 600 mg/kg. Doses of 300 mg/kg APAP or greater caused death in the homozygous knockout mice only, and those that survived showed a greater severity in hepatic damage than the wild-type mice, as demonstrated by increased plasma alanine aminotransferase activity, decreased hepatic non-protein sulfhydryl (NPSH) content, and centrilobular hepatocellular necrosis. The high sensitivity of Nrf2-deficient mice was confirmed from observations made at 0, 2, 8, and 24 h after dosing with 300 mg/kg APAP; increased anti-APAP immunoreactivity was also noted in their livers at 2 h. Untreated homozygous knockout mice showed both a lower UDP-GT activity and NPSH content, which corresponded to decreased mRNA levels of UDP-GT (Ugt1a6) and the heavy chain of gammaGCS, respectively. These results show that Nrf2 plays a protective role against APAP hepatotoxicity by regulating both drug metabolizing enzymes and antioxidant genes through the ARE.

Acetaminophen↗

PubMed2003

Epididymal phenotype in luteinizing hormone receptor knockout animals and its response to testosterone replacement therapy.

Previous studies reported that epididymis contains functional LH receptors. The LH receptor knockout mice, which have epididymal phenotypes, gave us an opportunity to test the hypothesis that testosterone replacement alone may not be sufficient to reverse phenotypes to wild-type epididymis. The morphological phenotype in knockout animals includes a decrease in luminal diameter of the proximal and distal caput and cauda epididymis, the absence of clear and halo cells in the epithelial lining, a decrease in the height of principal cells and the number of cells containing cilia, a decrease in cilia length, and a change from basal to central location of nuclei in the principal cells. The biochemical phenotype includes a decrease in periodic acid-Schiff reaction product, reflecting the glycogen and glycoprotein synthesis and secretion, a decrease in androgen receptor (AR) and estrogen receptor (ER)beta, and an increase in ERalpha levels. Twenty-one-day testosterone replacement therapy in 30-day-old knockout animals reversed some, but not all, morphological and biochemical phenotypes. Those that did not reverse include luminal diameters of proximal and distal caput and cauda epididymis, the percentage of ciliated principal cells in caput epididymis, and nuclear AR localization. In summary, while our results reaffirm that androgens are important for normal epididymal morphology and function, they indicate that LH could be required for certain facets of epididymal morphology and/or function.

Animals↗

PubMed2004

Inducible nitric oxide synthase mediates delayed cardioprotection induced by morphine in vivo: evidence from pharmacologic inhibition and gene-knockout mice.

BACKGROUND: It is not known whether morphine induces delayed cardioprotection against ischemia and reperfusion. The authors measured the delayed preconditioning induced by morphine and determined the role of inducible nitric oxide synthase (iNOS) in mediating this effect using a pharmacological inhibitor and iNOS gene-knockout mice. METHODS: Adult male wild-type and iNOS gene-knockout (B6, 129) mice were treated with morphine (0.3 or 0.1 mg/kg intraperitoneal) or saline. Twenty-four hours later, mice were subjected to 45 min of coronary artery occlusion followed by 120 min of reperfusion. S-methylthiourea sulfate (3 mg/kg, intraperitoneal) was given 30 min before the occlusion to block iNOS. Infarct size as a percentage of the area at risk was determined by triphenyltetrazolium chloride staining. iNOS and endothelial nitric oxide synthase expression were measured by Western blot. RESULTS: Infarct size was significantly reduced in wild-type mice from 43.1 +/- 5.3% in the saline group to 22.4 +/- 4.4% in the higher-dose morphine group (0.3 mg/kg) (P < 0.05). This cardioprotective effect was abolished by S-methylthiourea sulfate (43.3 +/- 3.9%) and was absent in iNOS gene-knockout mice (42.3 +/- 4.7%). Pretreatment with the lower dose of morphine (0.1 mg/kg) did not reduce infarct size (41.1 +/- 5.4%). A significant increase in myocardial iNOS expression was observed 24 h after morphine administration (0.3 mg/kg but not 0.1 mg/kg; P < 0.05), whereas endothelial nitric oxide synthase remained unchanged. CONCLUSIONS: : Pretreatment with morphine induces delayed cardioprotection in mice. The authors demonstrated an obligatory role for iNOS in mediating morphine-induced delayed cardioprotection.

Analgesics, Opioid↗