Consequences of haemolysis without haptoglobin

Haptoglobin (Hp), a conserved plasma glycoprotein, forms very stable soluble complexes with free plasma haemoglobin. Haemoglobin binding by haptoglobin is thought to be important in the rapid hepatic clearance of haemoglobin from the plasma and in the inhibition of glomerular filtration of haemoglobin. It is thought to reduce haemoglobin-induced renal damage during haemolysis. To evaluate these functions, Hp knockout (Hp-/-) mice were created. The Hp-/- mouse was generated by a standard gene replacement technique in mouse embryonic stem cells. These mice were evaluated with and without haemolysis using several parameters: mortality, haemoglobin clearance, renal tissue damage and function. Hp-/- mice were viable but had a small, significant reduction in postnatal viability. The lack of Hp did not impair clearance of free plasma haemoglobin. Induction of severe haemolysis by phenylhydrazine caused extensive haemoglobin precipitation in the renal tubular cells. However, haemoglobin precipitation in the kidney was not increased in Hp-/- mice. Nevertheless, Hp-/- mice were more susceptible to phenylhydrazine with a mortality rate of 55% in Hp-/- mice versus 18% in Hp+/+ mice. In general, phenylhydrazine-treated Hp-/- mice suffered greater tissue damage, as evidenced by the induction of a hepatic acute phase response, resulting in increased plasma alpha1-acidic glycoprotein (AGP) levels and higher plasma malonaldehyde (MDA) and 4-hydroxy-2(E)-nonenal (HNE) levels. Gross pathological analysis indicated that the kidney was the most affected tissue in phenylhydrazine-treated Hp-/- and Hp+/+ mice, and Hp-/- mice were more severely affected. They had lower mitotic indices in their kidneys, higher basal levels of renal lipid peroxidation, as evidenced by levels of malonaldehyde and 4-hydroxy-2(E)-nonenal (MDA/HNE) and elevated levels of 8-hydroxyguanine (but not other products of oxidative DNA damage). There also was increased induction of haem oxygenase-1. The more severe renal damage in Hp-/- mice was also evident in the delayed erythropoietin gene expression and poorer renal clearance of [3H]-inulin. The reduction in glomerular filtration function in Hp+/+ and Hp-/- mice could be restored to baseline by vasodilators (prazosin or diazoxide), implicating renal vasoconstriction as a major mechanism of acute renal failure during induced haemolysis. These data suggest that Hp plays a pivotal role in reducing renal oxidative damage during haemolysis.     

Haptoglobin and malaria

Abstract

Haptoglobin gene knockout mice and wild-type controls were infected with Plasmodium berghei ANKA or Plasmodium chabaudi. The peak parasitaemia and parasite burden were higher in Hp^-/- mice than in Hp^+/+ mice. The increase in spleen weight following malaria infection was smaller in Hp^-/- mice than in Hp^+/+ animals. The occurrence of cerebral malaria in P. berghei ANKA infection was not different in Hp gene knockout mice and their controls.     

The effect of spin traps on phenylhydrazine-induced haemolysis

Spin-trapping agents have been used to study the involvement of free radicals in phenylhydrazine-induced haemolysis. Spin traps were found to decrease the rate of oxygen uptake and the rate of haemoglobin oxidation in the reaction of phenylhydrazine with oxyhaemoglobin. Spin traps were also found to inhibit haemolysis and lipid peroxidation in phenylhydrazine-treated fresh human erythrocytes, with a concomitant production of phenyl radical spin adducts. Lipophilic spin traps were found to be more effective inhibitors of haemolysis than their hydrophilic analogues.     

Haptoglobin in lung defence

Abstract

Haptoglobin (Hp) has been known to be associated with the host defence response to infection and inflammation. The biological functions of Hp can be related to its ability to bind haemoglobin or to modulate immune response. Hp is expressed at a high level in lung cells, yet its protective role (s) in the lung is not known. Using transgenic mice overexpressing Hp, we demonstrated that Hp can reduce blood-induced lung injury. Hp-mediated haemoglobin catabolism in lung cells appears to be linked to iron mobilization, and may be an efficient mechanism to reduce oxidative damage associated with haemolysis.     

Irgm 1参与动脉粥样硬化斑块的形成

目的:探讨免疫相关GTP酶1(Irgm 1)对小鼠血管动脉粥样硬化(AS)斑块形成的影响。方法:高脂饲料喂养野生型(WT)、ApoE~(-/-)Irgm 1~(+/+)和ApoE~(-/-)Irgm1~(+/-)小鼠3个月,建立AS模型;取小鼠主动脉弓,免疫荧光染色方法观察WT和ApoE~(-/-)Irgm 1~(+/+)小鼠血管AS斑块中Irgm 1的表达情况及部位;Western blot方法检测WT和ApoE~(-/-)Irgm 1~(+/+)小鼠血管AS斑块中Irgm 1蛋白表达情况;Q-PCR方法检测WT和ApoE~(-/-)Irgm 1~(+/+)小鼠血管AS斑块中Irgm 1 m RNA表达情况;油红O染色观察ApoE~(-/-)Irgm1~(+/+)和ApoE~(-/-)Irgm1~(+/-)小鼠血管AS斑块形成情况;结果:与WT组相比,ApoE~(-/-)Irgm 1~(+/+)组小鼠主动脉弓AS斑块中Irgm 1+细胞明显增多,Irgm 1+细胞主要位于血管AS斑块的表面;与WT组相比,ApoE~(-/-)Irgm 1~(+/+)组小鼠血管AS斑块中Irgm 1蛋白表达显著增多(P0.001),Irgm 1 m RNA表达显著增多(P0.01);与ApoE~(-/-)Irgm1~(+/-)组相比,ApoE~(-/-)Irgm1~(+/+)组小鼠主动脉弓AS斑块面积显著增大(P0.01);结论:Irgm 1能够促进血管AS斑块的形成。     

Periostin Deficiency Increases Bone Damage and Impairs Injury Response to Fatigue Loading in Adult Mice

Bone damage removal and callus formation in response to fatigue loading are essential to prevent fractures. Periostin (Postn) is a matricellular protein that mediates adaptive response of cortical bone to loading. Whether and how periostin influences damage and the injury response to fatigue remains unknown. We investigated the skeletal response of Postn ^-/- and Postn ^+/+ mice after fatigue stimulus by axial compression of their tibia. In Postn ^+/+ mice, cracks number and surface (CsNb, CsS) increased 1h after fatigue, with a decrease in strength compared to non-fatigued tibia. At 15 days, CsNb had started to decline, while CtTV and CtBV increased in fatigued vs non-fatigued tibia, reflecting a woven bone response that was present in 75% of the fatigued bones. Cortical porosity and remodelling also prominently increased in the fatigued tibia of Postn ^+/+ mice. At 30 days, paralleling a continuous removal of cortical damage, strength of the fatigued tibia was similar to the non-fatigue tibia. In Postn ^-/- mice, cracks were detectable even in the absence of fatigue, while the amount of collagen crosslinks and tissue hardness was decreased compared to Postn ^+/+. Fatigue significantly increased CsNb and CsS in Postn ^-/-, but was not associated with changes in CtTV and CtBV, as only 16% of the fatigued bones formed some woven bone. Cortical porosity and remodelling did not increase either after fatigue in Postn ^-/- , and the level of damage remained high even after 30 days. As a result, strength remained compromised in Postn ^-/- mice. Contrary to Postn ^+/+ , which osteocytic lacunae showed a change in the degree of anisotropy (DA) after fatigue, Postn^-/- showed no DA change. Hence periostin appears to influence bone materials properties, damage accumulation and repair, including local modeling/remodeling processes in response to fatigue. These observations suggest that the level of periostin expression could influence the propensity to fatigue fractures.     

Perforin deficiency attenuates collagen-induced arthritis

Collagen-induced arthritis (CIA), an approved animal model for rheumatoid arthritis, is thought to be a T cell-dependent disease. There is evidence that CD8⁺ T cells are a major subset controlling the pathogenesis of CIA. They probably contribute to certain features of disease, namely tissue destruction and synovial hyperplasia. In this study we examined the role of perforin (pfp), a key molecule of the cytotoxic death pathway that is expressed mainly in CD8⁺ T cells, for the pathogenesis of CIA. We generated DBA/1J mice suffering from mutations of the pfp molecule, DBA/1J-pfp^-/-, and studied their susceptibility to arthritis. As a result, pfp-deficient mice showed a reduced incidence (DBA/1J-pfp^+/+, 64%; DBA/1J-pfp^-/-, 54%), a slightly delayed onset (onset of disease: DBA/1J-pfp^+/+, 53 ± 3.6; DBA/1J-pfp^-/-, 59 ± 4.9 (mean ± SEM), and milder form of the disease (maximum disease score: DBA/1J-pfp^+/+, 7.3 ± 1.1; DBA/1J-pfp^-/-, 3.4 ± 1.4 (mean ± SEM); P < 0.05). Concomitantly, peripheral T cell proliferation in response to the specific antigen bovine collagen II was increased in pfp^-/- mice compared with pfp^+/+ mice, arguing for an impaired killing of autoreactive T cells caused by pfp deficiency. Thus, pfp-mediated cytotoxicity is involved in the initiation of tissue damage in arthritis, but pfp-independent cytotoxic death pathways might also contribute to CIA.     

Plasminogen Deficiency Causes Reduced Corticospinal Axonal Plasticity and Functional Recovery after Stroke in Mice

Tissue plasminogen activator (tPA) has been implicated in neurite outgrowth and neurological recovery post stroke. tPA converts the zymogen plasminogen (Plg) into plasmin. In this study, using plasminogen knockout (Plg^-/-) mice and their Plg-native littermates (Plg^+/+), we investigated the role of Plg in axonal remodeling and neurological recovery after stroke. Plg^+/+ and Plg^-/- mice (n = 10/group) were subjected to permanent intraluminal monofilament middle cerebral artery occlusion (MCAo). A foot-fault test and a single pellet reaching test were performed prior to and on day 3 after stroke, and weekly thereafter to monitor functional deficit and recovery. Biotinylated dextran amine (BDA) was injected into the left motor cortex to anterogradely label the corticospinal tract (CST). Animals were euthanized 4 weeks after stroke. Neurite outgrowth was also measured in primary cultured cortical neurons harvested from Plg^+/+ and Plg^-/- embryos. In Plg^+/+ mice, the motor functional deficiency after stroke progressively recovered with time. In contrast, recovery in Plg^-/- mice was significantly impaired compared to Plg^+/+ mice (p<0.01). BDA-positive axonal density of the CST originating from the contralesional cortex in the denervated side of the cervical gray matter was significantly reduced in Plg^-/- mice compared with Plg^+/+ mice (p<0.05). The behavioral outcome was highly correlated with the midline-crossing CST axonal density (R²>0.82, p<0.01). Plg^-/- neurons exhibited significantly reduced neurite outgrowth. Our data suggest that plasminogen-dependent proteolysis has a beneficial effect during neurological recovery after stroke, at least in part, by promoting axonal remodeling in the denervated spinal cord.     

P2Y2R Deficiency Attenuates Experimental Autoimmune Uveitis Development

We aimed to study the role of the nucleotide receptor P2Y₂R in the development of experimental autoimmune uveitis (EAU). EAU was induced in P2Y₂^+/+ and P2Y₂^-/- mice by immunization with IRBP peptide or by adoptive transfer of in vitro restimulated semi-purified IRBP-specific enriched T lymphocytes from spleens and lymph nodes isolated from native C57Bl/6 or P2Y₂^+/+ and P2Y₂^-/- immunized mice. Clinical and histological scores were used to grade disease severity. Splenocytes and lymph node cell phenotypes were analyzed using flow cytometry. Semi-purified lymphocytes and MACS-purified CD4⁺ T lymphocytes from P2Y₂^+/+ and P2Y₂^-/- immunized mice were tested for proliferation and cytokine secretion. Our data show that clinical and histological scores were significantly decreased in IRBP-immunized P2Y₂^-/- mice as in P2Y₂^-/- mice adoptively transfered with enriched T lymphocytes from C57Bl/6 IRBP-immunized mice. In parallel, naïve C57Bl/6 mice adoptively transferred with T lymphocytes from P2Y₂^-/- IRBP-immunized mice also showed significantly less disease. No differences in term of spleen and lymph node cell recruitment or phenotype appeared between P2Y₂^-/- and P2Y₂^+/+ immunized mice. However, once restimulated in vitro with IRBP, P2Y₂^-/- T cells proliferate less and secrete less cytokines than the P2Y₂^+/+ one. We further found that antigen-presenting cells of P2Y₂^-/- immunized mice were responsible for this proliferation defect. Together our data show that P2Y₂^-/- mice are less susceptible to mount an autoimmune response against IRBP. Those results are in accordance with the danger model, which makes a link between autoreactive lymphocyte activation, cell migration and the release of danger signals such as extracellular nucleotides.     

TLR2 mediates immunity to experimental cysticercosis

Information concerning TLR-mediated antigen recognition and regulation of immune responses during helminth infections is scarce. TLR2 is a key molecule required for innate immunity and is involved in the recognition of a wide range of viruses, bacteria, fungi and parasites. Here, we evaluated the role of TLR2 in a Taenia crassiceps cysticercosis model. We compared the course of T. crassiceps infection in C57BL/6 TLR2 knockout mice (TLR2^-/-) with that in wild type C57BL/6 (TLR2^+/+) mice. In addition, we assessed serum antibody and cytokine profiles, splenic cellular responses and cytokine profiles and the recruitment of alternatively activated macrophages (AAMφs) to the site of the infection. Unlike wild type mice, TLR2^-/- mice failed to produce significant levels of inflammatory cytokines in either the serum or the spleen during the first two weeks of Taenia infection. TLR2^-/- mice developed a Th2-dominant immune response, whereas TLR2^+/+ mice developed a Th1-dominant immune response after Taenia infection. The insufficient production of inflammatory cytokines at early time points and the lack of Th1-dominant adaptive immunity in TLR2^-/- mice were associated with significantly elevated parasite burdens; in contrast, TLR2^+/+ mice were resistant to infection. Furthermore, increased recruitment of AAMφs expressing PD-L1, PD-L2, OX40L and mannose receptor was observed in TLR2^-/- mice. Collectively, these findings indicate that TLR2-dependent signaling pathways are involved in the recognition of T. crassiceps and in the subsequent activation of the innate immune system and production of inflammatory cytokines, which appear to be essential to limit infection during experimental cysticercosis.     

Lipoprotein X Causes Renal Disease in LCAT Deficiency

Human familial lecithin:cholesterol acyltransferase (LCAT) deficiency (FLD) is characterized by low HDL, accumulation of an abnormal cholesterol-rich multilamellar particle called lipoprotein-X (LpX) in plasma, and renal disease. The aim of our study was to determine if LpX is nephrotoxic and to gain insight into the pathogenesis of FLD renal disease. We administered a synthetic LpX, nearly identical to endogenous LpX in its physical, chemical and biologic characteristics, to wild-type and Lcat^-/- mice. Our in vitro and in vivo studies demonstrated an apoA-I and LCAT-dependent pathway for LpX conversion to HDL-like particles, which likely mediates normal plasma clearance of LpX. Plasma clearance of exogenous LpX was markedly delayed in Lcat^-/- mice, which have low HDL, but only minimal amounts of endogenous LpX and do not spontaneously develop renal disease. Chronically administered exogenous LpX deposited in all renal glomerular cellular and matrical compartments of Lcat^-/- mice, and induced proteinuria and nephrotoxic gene changes, as well as all of the hallmarks of FLD renal disease as assessed by histological, TEM, and SEM analyses. Extensive in vivo EM studies revealed LpX uptake by macropinocytosis into mouse glomerular endothelial cells, podocytes, and mesangial cells and delivery to lysosomes where it was degraded. Endocytosed LpX appeared to be degraded by both human podocyte and mesangial cell lysosomal PLA₂ and induced podocyte secretion of pro-inflammatory IL-6 in vitro and renal Cxl10 expression in Lcat^-/- mice. In conclusion, LpX is a nephrotoxic particle that in the absence of Lcat induces all of the histological and functional hallmarks of FLD and hence may serve as a biomarker for monitoring recombinant LCAT therapy. In addition, our studies suggest that LpX-induced loss of endothelial barrier function and release of cytokines by renal glomerular cells likely plays a role in the initiation and progression of FLD nephrosis.     

Alterations of red cell membranes from phenylhydrazine-treated rabbits

Reticulocytosis was induced in rabbits by two methods: phlebotomy and injection of phenylhydrazine. Normal erythrocytes, reticulocytes from bed rabbits, reticulocytes from phenylhydrazine-treated rabbits, and erythrocytes treated in vitro with phenylhydrazine were compared with respect to their plasma membrane labeling by galactose oxidase and NaB³H₄, and lactoperoxidase-catalyzed incorporation of ¹²⁵I. Normal erythrocyte membranes and membranes from reticulocytes of bled rabbits showed almost identical labeling patterns, the presence of 2–3 glycoproteins with moderate to low mobilities on dodecyl sulfate acrylamide gel electrophoresis. Labeling in the absence of enzyme was negligible. In contrast, the reticulocytes from phenylhydrazine-treated rabbits exhibited a large incorporation of tritium without prior treatment with galactose oxidase. Even after prereduction with unlabeled NaBH₄ to remove this nonspecific labeling, the labeled glycoprotein components found in normal erythrocytes were not detectable. Normal erythrocytes treated in vitro with phenylhydrazine, washed, and labeled with galactose oxidase had labeling patterns, including high nonspecific incorporation of ³H, similar to those observed with in vivo phenylhydrazine treatment.Solubilization of membranes with lithium diiososalicylate followed by partitioning with phenol showed that the same glycoproteins were presented in normal or phenylhydrazine membranes, although only the former extract was labeled by galactose oxidase. Individual carbohydrates from the membranes were analyzed by gas-liquid chromatography and, in the case of hexosamines, on the amino acid analyzer. The results of these analyses indicated a slight decline in galactose content with phenylhydrazine treatment. Reticulocyte membranes from bled rabbits also showed a decrease in galactose content, although it was less pronounced.Most of the label incorporated by nonspecific borohydride labeling of membranes from phenylhydrazine-treated animals was found associated with protein. The modified amino acids from labeled proteins are similar to those formed in reactions of oxidized lipids and proteins in model systems.     

Haptoglobin in lung defence

Haptoglobin (Hp) has been known to be associated with the host defence response to infection and inflammation. The biological functions of Hp can be related to its ability to bind haemoglobin or to modulate immune response. Hp is expressed at a high level in lung cells, yet its protective role(s) in the lung is not known. Using transgenic mice overexpressing Hp, we demonstrated that Hp can reduce blood-induced lung injury. Hp-mediated haemoglobin catabolism in lung cells appears to be linked to iron mobilization, and may be an efficient mechanism to reduce oxidative damage associated with haemolysis.     

Adiponectin Receptor 2 Deficiency Results in Reduced Atherosclerosis in the Brachiocephalic Artery in Apolipoprotein E Deficient Mice

Adiponectin has been shown to have beneficial cardiovascular effects and to signal through the adiponectin receptors, AdipoR1 and AdipoR2. The original aim of this study was to investigate the effect of combined AdipoR1 and AdipoR2 deficiency (AdipoR1^-/-AdipoR2^-/-) on atherosclerosis. However, we made the interesting observation that AdipoR1 ^-/- AdipoR2 ^-/- leads to embryonic lethality demonstrating the critical importance of the adiponectin signalling system during development. We then investigated the effect of AdipoR2-ablation on the progression of atherosclerosis in apolipoprotein E deficient (ApoE ^-/-) mice. AdipoR2^-/-ApoE^-/- mice fed an atherogenic diet had decreased plaque area in the brachiocephalic artery compared with AdipoR2 ^+/+ApoE^-/- littermate controls as visualized in vivo using an ultrasound biomicroscope and confirmed by histological analyses. The decreased plaque area in the brachiocephalic artery could not be explained by plasma cholesterol levels or inflammatory status. However, accumulation of neutral lipids was decreased in peritoneal macrophages from AdipoR2^-/-ApoE^-/- mice after incubation with oxidized LDL. This effect was associated with lower CD36 and higher ABCA1 mRNA levels in peritoneal macrophages from AdipoR2^-/-ApoE^-/- mice compared with AdipoR2^+/+ApoE^-/- controls after incubation with oxidized LDL. In summary, we show that adiponectin receptors are crucial during embryonic development and that AdipoR2-deficiency slows down the progression of atherosclerosis in the brachiocephalic artery of ApoE-deficient mice.     

α-Defensins Induce a Post-translational Modification of Low Density Lipoprotein (LDL) That Promotes Atherosclerosis at Normal Levels of Plasma Cholesterol

Approximately one-half of the patients who develop clinical atherosclerosis have normal or only modest elevations in plasma lipids, indicating that additional mechanisms contribute to pathogenesis. In view of increasing evidence that inflammation contributes to atherogenesis, we studied the effect of human neutrophil α-defensins on low density lipoprotein (LDL) trafficking, metabolism, vascular deposition, and atherogenesis using transgenic mice expressing human α-defensins in their polymorphonuclear leukocytes (Def^+/+). Accelerated Def^+/+ mice developed α-defensin·LDL complexes that accelerate the clearance of LDL from the circulation accompanied by enhanced vascular deposition and retention of LDL, induction of endothelial cathepsins, increased endothelial permeability to LDL, and the development of lipid streaks in the aortic roots when fed a regular diet and at normal plasma levels of LDL. Transplantation of bone marrow from Def^+/+ to WT mice increased LDL clearance, increased vascular permeability, and increased vascular deposition of LDL, whereas transplantation of WT bone marrow to Def^+/+ mice prevented these outcomes. The same outcome was obtained by treating Def^+/+ mice with colchicine to inhibit the release of α-defensins. These studies identify a potential new link between inflammation and the development of atherosclerosis.     

Analysis of the polymorphism of haptoglobins in Sardinia

The present study reports an analysis of the distribution of Hp polymorphism in Sardinia carried out on a sample of 871 individuals. The samples are examined according to the division by provinces and by historical-geographical regions. In both cases the phenotype Hp 2-1 has high frequency and a North-South trend. The Hp^*1 is the most common gene in every part of Sardinia. In general the distribution of Hp in Sardinia is not significantly heterogeneous in our sample, although presenting a discreet variability. The synthesis of our data and of the data available in literature shows some historical-geographicalareas, especially in the central north of the island, with particularly high Hp^*1 frequency (Baronie Siniscola and Orosei, Barbagie Ollorai and Belvì and Planargia; Sarrabus in the South). Similar high values of Hp^*1 have been highlighted in some groups of population in the Mediterranean basin, characterized by a certain geographical and/or cultural isolation.     

Phenotypic analysis of boldit Galr1 knockout mice reveals a role for GALR1 galanin receptor in modulating seizure activity but not nerve regeneration

The GALR1 galanin receptor is expressed at high levels withinthe central nervous system and is hypothesised to play asignificant role in many of the central actions of galanin. Todetermine which specific actions of galanin are mediated byGALR1, we have developed mice that carry an insertionalinactivating mutation within the first coding exon of the geneencoding GALR1 (Galr1). HomozygousGalr1 ^-/-mice are viable. Both male and female mice exhibit reducedcirculating levels of insulin-like growth factor-I (IGF-I) butno significant difference in growth rate relative to Galr1 ^+/+ controls. Female homozygousGalr1 ^-/-mice are capable of breeding and nursing offspring. Functionalrecovery after sciatic nerve crush is not significantlydifferent in Galr1 ^-/- mice relative to Galr1 ^+/+ controls, indicating that GALR1 does not mediate the nerve regenerative effects of galanin. However, homozygous Galr1 ^-/- mice exhibit spontaneous seizures, identifying a critical role for GALR1 in mediating the anti-seizure activity of galanin.     

Loss of NAC1 Expression Is Associated with Defective Bony Patterning in the Murine Vertebral Axis

NAC1 encoded by NACC1 is a member of the BTB/POZ family of proteins and participates in several pathobiological processes. However, its function during tissue development has not been elucidated. In this study, we compared homozygous null mutant Nacc1^-/- and wild type Nacc1^+/+ mice to determine the consequences of diminished NAC1 expression. The most remarkable change in Nacc1^-/- mice was a vertebral patterning defect in which most knockout animals exhibited a morphological transformation of the sixth lumbar vertebra (L6) into a sacral identity; thus, the total number of pre-sacral vertebrae was decreased by one (to 25) in Nacc1^-/- mice. Heterozygous Nacc1^+/- mice had an increased tendency to adopt an intermediate phenotype in which L6 underwent partial sacralization. Nacc1^-/- mice also exhibited non-closure of the dorsal aspects of thoracic vertebrae T10-T12. Chondrocytes from Nacc1^+/+ mice expressed abundant NAC1 while Nacc1^-/- chondrocytes had undetectable levels. Loss of NAC1 in Nacc1^-/- mice was associated with significantly reduced chondrocyte migratory potential as well as decreased expression of matrilin-3 and matrilin-4, two cartilage-associated extracellular matrix proteins with roles in the development and homeostasis of cartilage and bone. These data suggest that NAC1 participates in the motility and differentiation of developing chondrocytes and cartilaginous tissues, and its expression is necessary to maintain normal axial patterning of murine skeleton.     

New immunological investigations on Helicobacter pylori‐induced gastric ulcer in patients

Although Helicobacter pylori (Hp) plays an important role in the pathogenesis of chronic gastritis and gastric ulcer, little is known about the probable mechanisms of these types of gastrointestinal damage. To determine the precise mechanisms involved in ulcer formation, immune responses in patients with gastric ulcer (GUP) caused by Hp infection (Hp⁺) were compared with those of other gastritis patients (GP). The sensitivity and proliferation of peripheral blood mononuclear cells (PBMNCs) obtained from patients were evaluated by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT) assay against exposure with complex Hp crude antigen (HPCA) and mitogen (phytohemagglutinin, PHA). Production of inflammatory cytokines, including interleukin (IL)‐1β and IL‐8, in serum and supernatants of PBMNCs were then measured by ELISA. It was found that, after stimulation with PHA, both IL‐8 and IL‐1β concentrations in sera and supernatants as well as proliferation and sensitivity were statistically greater in GUP Hp⁺ than GP Hp^?. Furthermore, HPCA inhibited the proliferation of PBMNCs dose‐dependently; however, it stimulated IL‐8 and IL‐1β production in supernatants of mononuclear cells. Therefore, the up‐regulated concentrations of IL‐8 and IL‐1β may have been caused by increase in the size of mononuclear cell subpopulations or in their cytokine secretory activity, indicating the greatest cell responsiveness in GUP Hp⁺ patients. These results suggest that tissue damage and ulcers occur in patients who produce more IL‐8 and IL‐1β than patients who do not develop ulcers; the former consequently have more activated immune cells at the site of infection. Therefore, both host responses and Hp virulence factors may be involved in the development of gastric ulcers.