The renoprotective effects of taurine against diabetic nephropathy via the p38 MAPK and TGF-ß/Smad2/3 signaling pathways.

Diabetic nephropathy (DN), a severe diabetes complication, causes kidney morphological and structural changes due to extracellular matrix accumulation. This accumulation is caused mainly by oxidative stress. Semi-essential amino acid derivative taurine has powerful antioxidant and antifibrotic effects. The aim of this study was to investigate the renoprotective effects of taurine through its possible roles in oxidative stress, extracellular matrix proteins, and the signaling pathways associated with the accumulation of extracellular matrix proteins in DN rats. 29 Wistar albino rats were randomly separated into control, taurine, diabetes, and diabetes + taurine groups. Diabetes animals were injected 45 mg/kg streptozosine. Taurine is given by adding to drinking water as 1% (w/v). Urine, serum, and kidney tissue were collected from rats for biochemical and histological analysis after 12 weeks. According to the studies, taurine significantly reduces the levels of malondialdehyde (MDA), total oxidant status (TOS), and protein expression of NADPH oxidase 4 (NOX4) that increase in diabetic kidney tissue. Also, decreased superoxide dismutase (SOD) activity levels significantly increased with taurine in diabetic rats. Moreover, increased mRNA and protein levels of fibronectin decreased with taurine. The matrix metalloproteinase (MMP)-2 and MMP-9 activities and their mRNA levels increased significantly, and this increase was significantly summed with taurine. There was a decrease in mRNA expression of Extracellular matrix metalloproteinase inducer (EMMPRIN). Taurine significantly increased this decrease. Diabetes increased mRNA expressions of transforming growth factor (TGF)-ß and Smad2/3. Taurine significantly reduced this induction. TGF-ß protein expression, p38, and Smad2/3 activations were also inhibited, but taurine was suppressed significantly. All these findings indicate that taurine may be an effective practical strategy to prevent renal diabetic injury.

Role of p38 MAPK, Akt and NFκB in renoprotective effects of nebivolol on renal ischemia-reperfusion injury in rats.

Renal ischemia-reperfusion (I-R) injury is a complex pathophysiologic condition characterized by oxidative stress, inflammation and apoptosis. We investigated the potential renoprotective effect of nebivolol, a ß1 adrenergic receptor blocker, against renal I-R injury. We focused on the role of nebivolol in activating p38 mitogen-activated protein kinase (MAPK) signaling, Akt (protein kinase B) and nuclear factor-κB (NFκB) transcription factors, which contribute to oxidative stress, inflammation and apoptosis during renal I-R. We divided 20 adult male Wistar albino rats into three experimental groups. Group 1 was a sham control in which only laparotomy was performed. Group 2 was the I-R group in which both kidneys were made ischemic for 45 min, then reperfused for 24 h. Group 3 was the I-R + nebivolol group in which 10 mg/kg nebivolol was administrated by gavage for 7 days before I-R. We measured Inflammation, oxidative stress and active caspase-3 as well as activation of p38 MAPK, Akt (protein kinase B) and NFκB transcription factor. Nebivolol significantly reduced oxidative stress and increased superoxide dismutase levels during renal I-R. We found that nebivolol significantly decreased interstitial inflammation, and TNF-α and interleukin-1ß mRNA expression. Nebivolol significantly reduced active caspase-3 and kidney injury molecule-1 (KIM-1) expressions. Nebivolol also significantly decreased activation of p38 MAPK signaling and NFκB, and induced Akt activation during renal I-R. Our findings suggest that nebivolol may be useful for management of renal I-R injury.

A New Perspective on the Pathogenesis of Infantile Colic: Is Infantile Colic a Biorhythm Disorder?

OBJECTIVES: In this study, we investigated the relationship between infantile colic, migraine, and biorhythm regulation, by evaluating biochemical and molecular parameters. STUDY DESIGN: Healthy infants with and without infantile colic were eligible for this prospective cohort study. A questionnaire was applied. Between the 6th and 8th postnatal weeks, day and night circadian histone gene H3f3b mRNA expression and spot urine excretion of serotonin, cortisol, and 6-sulphatoxymelatonin were analyzed. RESULTS: Among the 95 infants included, 49 were diagnosed with infantile colic. In the colic group, defecation difficulty, sensitivity to light/sound, and maternal migraine frequency increased and sleep disruption was typical. In the melatonin analysis, the difference between day and night levels was significant in the control group, indicating an established circadian rhythm ( P = 0.014). In the colic group, there was no day-night difference ( P = 0.216) in melatonin, but serotonin levels were higher at night. In the cortisol analysis, day-night values were similar in both groups. Day-night variability of H3f3b mRNA levels between the groups was significant, indicating circadian rhythm disturbance in the colic group compared to the control group ( P = 0.003). Fluctuations in circadian genes and hormones expected in healthy rhythm were revealed in the control group, but were missing in the colic group. CONCLUSION: Due to the gaps in the etipathogenesis in infantile colic, a unique effective agent has not been discovered so far. This study, which demonstrated for the first time that infantile colic is a biorhythm disorder using molecular methods, fills the gap in this regard and points to a completely different perspective in terms of treatment.

Effect of ferulic acid on testicular damage caused by torsion-detorsion in rats.

Testicular torsion is twisting of the spermatic cord around its axis, which impairs blood flow and causes ischemia and formation of free radicals. Ferulic acid is a phenolic acid of the hydroxycinnamic family that is found in the seeds and leaves of plants; it is present in substantial amounts in fruits and vegetables. We investigated the protective effect of ferulic acid on experimental testicular torsion in rats. Animals were divided randomly into five groups: control, ethyl alcohol, torsion, torsion-detorsion, and torsion-detorsion + ferulic acid. Histopathology was assessed using hematoxylin and eosin, and periodic acid-Schiff staining. Tissues were assessed using TUNEL, active caspase-3, myeloperoxidase and inducible nitric oxide synthase immunostaining. Biochemical changes were assessed using assays for superoxide dismutase, malondialdehyde, glutathione peroxidase and glutathione. Ferulic acid reduced the levels of free radicals and increased the levels of antioxidants. Ferulic acid also reduced histopathological changes and germ cell differentiation in the testis following torsion-detorsion. Ferulic acid should be investigated further as a potential treatment for sequelae of torsion-detorsion injury.

Protective effects of alpha-lipoic acid on bleomycin-induced skin fibrosis through the repression of NADPH Oxidase 4 and TGF-ß1/Smad3 signaling pathways.

The aim of this study was to determine the protective effects of alpha-lipoic acid (ALA), which is known as a powerful antioxidant, and the possible related molecular mechanisms that mediate its favorable action on skin fibrosis in the bleomycin (BLM)-induced scleroderma (SSc) model in mice. The experimental design was established with four groups of eight mice: Control, ALA (100 mg/kg), BLM (5 µg/kg), and BLM + ALA group. BLM was administered via subcutaneous (sc) once a day while ALA was injected intraperitoneally (ip) twice a week for 21 days. Histopathological and biochemical analyses showed that ALA significantly reduced BLM-induced dermal thickness, inflammation score, and mRNA expression of tumor necrosis factor-alpha (TNF-α) in the skin. Besides, the mRNA expressions of the subunits of NADPH oxidase, which are Nox4 and p22phox, were found to be significantly induced in the BLM group. However, ALA significantly reduced their mRNA expression, which were in parallel to its decreasing effect on serum total oxidant status (TOS) level. Moreover, it was found that ALA downregulated the mRNA expressions of alpha-smooth muscle actin (α-SMA), collagen type I and fibronectin in the skin tissue of the BLM group. Additionally, it was shown that ALA reduced significantly the TGF-ß1 and p-Smad3 protein expressions in the BLM + ALA group. On the other hand, ALA did not exhibit any significant effect on the p38 mitogen-activated kinase (MAPK) activation induced by BLM. All these findings point out that ALA may be a promising treatment for the attenuation of skin fibrosis in SSc patients.

The effects of hypothyroidism and hyperthyroidism on placental Hofbauer cells of pregnant rats.

We investigated the effects of maternal thyroid disorders on Hofbauer cells of both the placenta and the fetus in pregnant rats. We divided 21 rats into three groups: control group, induced hypothyroidism (hypo) group and induced hyperthyroidism (hyper) group. Hypothyroidism was induced using propylthiouracil and hyperthyroidism was induced using L-thyroxine. We measured maternal weight, maternal free thyroxine, fetal weight, fetal viability and placental morphology. At the end of the experiment, fetuses of the hypo and hyper groups were less developed than those of the control group. In the hypo and hyper groups, the thickness of the labyrinth zone was decreased, but thickness of the basal zone and decidua basalis was increased. The number of Hofbauer cells was increased in both the hypo and hyper groups. Vascular endothelial growth factor expression was increased in both the hypo and hyper groups compared to controls. Our findings indicate that maternal thyroid disorders exert a negative effect on fetal growth and placental development.

The role of small GTPase Rac1 in ionizing radiation-induced testicular damage.

PURPOSE: The main acute and late effects of ionizing radiation on living organisms are the formation of reactive oxygen species (ROS), apoptosis and DNA damage. Since the Rac1 molecule is a subunit of the NADPH oxidase enzyme, it is known to participate in the generation of ROS. The aim of this study was to investigate the role of Rac1 molecule in testicular damage induced by low (0.02 Gy), medium (0.1 Gy) and high (5 Gy) dose irradiation. MATERIAL AND METHOD: In this study, Wistar rats (except the control group) were received whole body X-ray irradiation. Testicular tissues were removed 2 hours, 24 hours and 7 days after radiation exposure. Testicular damage was examined by hematoxylin-eosin staining and Johnsen's score. Immunohistochemical staining and G-LISA method were used to determine Rac1 expression and activation. To evaluate the generation of ROS in the testicular tissues, intracellular ROS, superoxide dismutase (SOD) and malondialdehyde (MDA) levels were measured. RESULTS: Increases in testicular damage were detected in all radiation exposed groups in a dose- and time-dependent manner. Compared to the control group, Rac1 expression decreased in all irradiated groups, while Rac1 activation increased. In addition, intracellular ROS and MDA levels were increased and SOD activity levels decreased in the irradiated groups compared to the control group. CONCLUSION: Our findings suggest that Rac1 has a role in the increase of intracellular ROS and lipid peroxidation which led to an increase in radiation- induced testicular damage.

Alpha-lipoic acid alleviates colistin nephrotoxicity in rats.

Colistin methanesulfonate (CMS), a clinical form of colistin, is widely used as a last-line treatment for multidrug-resistant (MDR) gram-negative bacterial infections in critically ill patients presenting a considerably high mortality rate. However, nephrotoxicity is considered to be a critical adverse effect that limits CMS's clinical use. Alpha-lipoic acid (ALA) is a strong antioxidant that is effective in preventing nephrotoxicity in many models. The aim of this study was to investigate ALA's ability to protect against nephrotoxicity induced by colistin in rats. Male Wistar albino rats were randomly divided into four groups. Group 1 was the control group (Control; n = 6), in which isotonic saline was administered to the rats. Group 2 was the ALA group (ALA; n = 6) in which rats received 100 mg/kg ALA. Groups 3 was the CMS (CMS; n = 7) in which 450.000 IU/kg/day of CMS was administered to the rats. Groups 4 was the CMS + ALA group (n = 6), in which rats were injected with 100 mg/kg of ALA 30 min before administration of CMS. All injections were performed intraperitoneally at 1, 4, 7, and 10 days. Urine was collected by using a metabolic cage for 24 h after each administration. The rats were euthanized under ether anesthesia after 24 h of the last administration. Blood and kidney samples then were collected for histological and biochemical analysis. ALA pretreatment could reverse the effects of colistin-induced nephrotoxicity, partly through its suppressing effect on Nox4 and caspase-3, which in turn results in its antioxidant and antiapoptotic effect. Therefore, ALA may be an effective strategy for the management of colistin nephrotoxicity.

Alpha lipoic acid attenuates iron induced oxidative acute kidney injury in rats.

Iron has been implicated in oxidative tissue injury owing to its ability to generate reactive oxygen species (ROS). We investigated the reno-protective effects of alpha lipoic acid (ALA) by investigating its effects on the kidney isoform of NADPH oxidase (Nox4) and the specific signaling pathways, p38 MAPK and PI3K/Akt, which participate in apoptosis and survival, respectively. We established four groups of seven rats: control, 100 mg/kg ALA, 80 mg/kg iron sucrose (IS) and IS + ALA. IS and ALA were injected intravenously and rats were sacrificied after 6 h. The mRNA expression of the subunits of NADPH oxidase, Nox4 and p22phox; tumor necrosis factor-alpha (TNF-α); and kidney injury molecule-1 (KIM-1) were measured using quantitative real time polymerase chain reaction (qRT-PCR). Active caspase-3 protein expression was evaluated by immunostaining. Also, p38 MAPK and PI3K/Akt signaling pathways were analyzed using western blot. ALA suppressed the mRNA expression of Nox4, p22phox, TNF-α and KIM-1. Active caspase-3 protein expression induced by IS was decreased by ALA. ALA also suppressed p38 MAPK and activated the PI3K/Akt signaling pathway following IS administration. We found that ALA may be an effective strategy for preventing oxidative acute kidney injury caused by IS.

Renoprotective Effects of Alpha Lipoic Acid on Iron Overload-Induced Kidney Injury in Rats by Suppressing NADPH Oxidase 4 and p38 MAPK Signaling.

We aimed to investigate the protective effect of alpha lipoic acid (ALA), a powerful antioxidant, against oxidative kidney damage induced by iron overload in rats. Male Wistar albino rats were separated into groups: control (n = 7), ALA (100 mg/kg (n = 7), iron sucrose (IS) (40 mg/kg) (n = 7), and IS + ALA (40 mg/kg IS administration followed by 100 mg/kg ALA) (n = 7). IS and ALA were injected weekly for 4 weeks via the tail vein. Blood and kidneys were collected at sacrification on day 29. Serum creatinine and iron levels were analyzed. Tubular injury and iron deposits were evaluated histopathologically. Additionally, iron, malondialdehyde (MDA), superoxide dismutase (SOD), catalase, and glutathione (GSH) levels and mRNA expressions of the subunits of NADPH oxidase, known as NOX4 and p22phox, tumor necrosis factor (TNF)-α, kidney injury molecule-1 (KIM-1), and also p38 MAPK signaling in the kidneys, were evaluated biochemically. In the IS group, serum creatinine and iron level, tubular dilation, and loss of brush border in the kidneys were significantly higher than those of the control. Although those changes were reduced by ALA, the differences were not statistically significant. However, ALA reduced significantly MDA level and increased SOD activity in the kidney during IS administration. ALA also significantly reduced mRNA expressions of NOX4 and p22phox induced by IS, which was parallel to significant decreases of TNF-α and KIM-1 mRNA expressions. Moreover, ALA could suppress the activation of p38 MAPK during IS administration. In conclusion, ALA may be an effective strategy to attenuate in IS-induced oxidative kidney injury.

Rat models of colistin nephrotoxicity: previous experimental researches and future perspectives.

Colistin is an old antibiotic, which is abandoned decades ago because of high nephrotoxicity rates. However, it is reintroduced to clinical medicine due to lack of newly discovered antibiotics and is still widely used for the treatment of resistant gram-negative infections. Discovering mechanisms to reduce nephrotoxicity risk is of significant importance since exposed patients may have many other factors that alter kidney functions. Several agents were evaluated in animal models of colistin nephrotoxicity as a means to prevent kidney injury. Considerable heterogeneity exists in terms of reporting colistin dosing and experimental designs. This issue leads clinicians to face difficulties in designing studies and sometimes may lead to report dosing strategies inadequately. Here, we present a review according to animal models of colistin nephrotoxicity using data gathered from previous experiments to draw attention on possible complexities that researchers may encounter.

Pretreatment with paricalcitol attenuates level and expression of matrix metalloproteinases in a rat model of renal ischemia-reperfusion injury
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BACKGROUND: Ischemia-reperfusion injury (IRI) is a leading cause of acute kidney injury (AKI). The inflammatory response that drives IRI involves upregulation of matrix metalloproteinases (MMPs), which results in proteolytic degradation of renal microvascular matrix. Evidence suggests a potential protective role of active vitamin D on ischemic injury by downregulating MMPs. In the present study, we aimed to determine the expression and level of MMP-2 and MMP-9 in renal IRI model and the potential beneficial effect of paricalcitol on both level and expression of MMPs and tubular injury caused by IRI. MATERIALS AND METHODS: 20 Wistar albino rats were divided into three groups: sham-operated, ischemia-reperfusion, and paricalcitol-pretreated. IRI model was induced by bilateral clamping of renal arteries for 45 minutes followed by 24 hours of reperfusion. The analysis of serum creatinine and levels of MMPs were performed after 24 hours of IRI. The effects of paricalcitol on the quantity and expression of MMP-2 and MMP-9 in renal tubular epithelial cells were investigated by enzyme-linked immunosorbent assay and immunohistochemistry, respectively. The pathological examinations were performed to score tubular damage by light microscopy. RESULTS: Creatinine levels decreased in the paricalcitol group, although this was not proven to be significant. Rats in the paricalcitol group showed significant decrease in both level and expression of MMPs and in tubular injury scores as compared to the IRI group. CONCLUSION: Paricalcitol may attenuate renal tubular injury caused by IRI by decreasing both level and expression of MMPs. Further studies are required to investigate the interplay between activated vitamin D and MMPs in AKI.
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Pretreatment with nebivolol attenuates level and expression of matrix metalloproteinases in a rat model of renal ischaemia-reperfusion injury.

AIM: Matrix metalloproteinases (MMPs) are zinc-containing proteinases that are involved in the degradation of extracellular matrix (ECM) and a number of cell surface proteins in order to maintain tissue homeostasis. They are involved in pathogenesis of several ischaemic organ injuries. In the present study, we aimed to determine the expression and level of MMP-2 and MMP-9 in renal ischaemia-reperfusion injury (IRI) model and the potential beneficial effect of nebivolol, a ß1 -adrenergic receptor blocker, on both MMP-2 and -9 level and expression and tubular injury caused by IRI. METHODS: Twenty Wistar albino rats were divided into three groups: sham-operated , ischaemia-reperfusion, and nebivolol-pretreated. IRI model was induced by bilateral clamping of renal arteries for 45 min followed by 24 h of reperfusion. The analysis of serum creatinine levels, quantity and expression of MMP-2 and MMP-9 were performed after 24 h of IRI. The effects of nebivolol on level and expression of MMP-2 and MMP-9 levels were investigated by enzyme-linked immunosorbent assay and immunohistochemistry, respectively. The pathological examinations were performed to score tubular damage by light microscopy. RESULTS: Creatinine levels increased significantly in the ischaemia-reperfusion group compared to the sham-operated group. Rats in the nebivolol-pretreated group showed significant decrease in expression and quantity of MMP-2 and MMP-9 during IRI. The pathological examinations demonstrated significantly low level of tubular injury score in nebivolol-pretreated group. CONCLUSION: Nebivolol attenuated IRI by decreasing the expression and level of MMP-2 and MMP-9.

The effects of α-lipoic acid against testicular ischemia-reperfusion injury in Rats.

Testicular torsion is one of the urologic emergencies occurring frequently in neonatal and adolescent period. Testis is sensitive to ischemia-reperfusion injury, and, therefore, ischemia and consecutive reperfusion cause an enhanced formation of reactive oxygen species that result in testicular cell damage and apoptosis. α-lipoic acid is a free radical scavenger and a biological antioxidant. It is widely used in the prevention of oxidative stress and cellular damage. We aimed to investigate the protective effect of α-lipoic acid on testicular damage in rats subjected to testicular ischemia-reperfusion injury. 35 rats were randomly divided into 5 groups: control, sham operated, ischemia, ischemia-reperfusion, and ischemia-reperfusion +lipoic acid groups, 2 h torsion and 2 h detorsion of the testis were performed. Testicular cell damage was examined by H-E staining. TUNEL and active caspase-3 immunostaining were used to detect germ cell apoptosis. GPx , SOD activity, and MDA levels were evaluated. Histological evaluation showed that α-lipoic acid pretreatment reduced testicular cell damage and decreased TUNEL and caspase-3-positive cells. Additionally, α-lipoic acid administration decreased the GPx and SOD activity and increased the MDA levels. The present results suggest that LA is a potentially beneficial agent in protecting testicular I/R in rats.

Resveratrol reduces matrix metalloproteinase-2 activity induced by oxygen-glucose deprivation and reoxygenation in human cerebral microvascular endothelial cells.

The main pathophysiology in cerebral ischemia is the structural alteration in the neurovascular unit, coinciding with neurovascular matrix degradation. Among the human matrix metalloproteinases (MMPs), MMP-2 and -9, known as gelatinases, are the key enzymes for degrading type IV collagen, which is the major component of the basal membrane that surrounds the cerebral blood vessel. In the present study, we investigated the effects of resveratrol on cytotoxicity, reactive oxygen species (ROS), and gelatinases (MMP-2 and -9) in human cerebral microvascular endothelial cells exposed to 6 hours of oxygen-glucose deprivation and a subsequent 24 hours of reoxygenation with glucose (OGD/R), to mimic ischemia/reperfusion in vivo. Lactate dehydrogenase increased significantly, in comparison to that in the normoxia group. ROS was markedly increased in the OGD/R group, compared to normoxia. Correspondingly, ROS was significantly reduced with 50 µM of resveratrol. The proMMP-2 activity in the OGD/R group showed a statistically significant increase from the control cells. Resveratrol preconditioning decreased significantly the proMMP-2 in the cells exposed to OGD/R in comparison to that in the OGD/R group. Our results indicate that resveratrol regulates MMP-2 activity induced by OGD/R via its antioxidant effect, implying a possible mechanism related to the neuroprotective effect of resveratrol.

Pioglitazone reduces peritoneal fibrosis via inhibition of TGF-ß, MMP-2, and MMP-9 in a model of encapsulating peritoneal sclerosis.

OBJECTIVE: Matrix metalloproteinases (MMPs) and transforming growth factor beta (TGF-ß) were increased in peritoneal dialysis patients with encapsulating peritoneal sclerosis (EPS) and in chlorhexidine gluconate (CG)-induced peritoneal sclerosing animal models. Peroxisome proliferator-activated receptors (PPARs) are the major regulators of key metabolic pathways of various inflammatory responses in fibrosing processes in most tissues. The objective of this study was to investigate the effect of pioglitazone (Pio), a synthetic PPAR-γ ligand, on the development of peritoneal fibrosis in CG-induced EPS rats. METHODS: Thirty-two Wistar albino rats were intraperitoneally injected with saline (C group n = 8) or with CG (1.5 mL/100 g; CG group, n = 8). Pio (30 mg/kg/day) was administered orally to another group of CG injected rats (the CG + Pio group, n = 8) and to another control group (Pio group, n = 8) from initiation to the end of this study. After 14 days of Pio administration, the rats were killed and the parietal and visceral peritoneum were harvested. TGF-ß, MMP-2, MMP-9, tissue inhibitor of metalloproteinases (TIMP)-1, and TIMP-2 activity assays and a morphological examination of the peritoneal tissues were performed. RESULTS: Pio significantly inhibited thickening of the submesothelial layer, fibrosis, and inflammation in the peritoneum. It also prevented increases in pro-MMP-2, pro-MMP-9, TIMP-1, and TGF-ß activities. CONCLUSION: Pio, via MMP and TGF-ß inhibition, may lessen accumulation of peritoneal extracellular matrix and fibrosis to some extent in an EPS model and might be a new approach to the amelioration of EPS.

Special-study modules in a problem-based learning medical curriculum: an innovative laboratory research practice supporting introduction to research methodology in the undergraduate curriculum.

We describe the organization of wet-lab special-study modules (SSMs) in the Central Research Laboratory of Dokuz Eylül Medical School, Izmir, Turkey with the aim of discussing the scientific, laboratory, and pedagogical aspects of this educational activity. A general introduction to the planning and functioning of these SSMs is given, along with specific examples. The wet-lab SSMs incorporate several innovative pedagogies: problem-based learning, research-based learning, practical laboratory education, team-based learning, and project-based learning. Oral and written evaluations show that the students find this activity rewarding. The wet-lab SSM model applied in the Research-Lab of Dokuz Eylül School of Medicine represents a format which is effective in training the students in research methodology, practical laboratory work, and independent learning.

Decrease in cell proliferation by an matrix metalloproteinase inhibitor, doxycycline, in a model of immune-complex nephritis.

AIM: Renal expression of matrix metalloproteinases (MMP) and tissue inhibitors of MMP (TIMP) contribute to the development of tubulointerstitial fibrosis characteristic of progressive forms of primary glomerulonephritis (GN). The aim of this study was to investigate the therapeutic effect of MMP inhibitor, doxycycline, administration in an experimental rat model of immune-complex nephritis (ICN). METHODS: The induction of immune-complex glomerulonephritis was carried out by the administration of an i.v. dose of 2 mg bovine serum albumin (BSA) daily for 28 days after 8 weeks of s.c. immunization with 1 mg of BSA in complete Freund's adjuvant. Doxycycline (30 mg/kg) was given daily (in groups 2 and 4) by gavage for 28 days. RESULTS: Animals treated with doxycycline showed significant reduction in glomerular area and cell proliferation than non-treated controls. Glomerular deposition of immunoglobulin (Ig)G and C3 was less intense in treated rats than non-treated controls. Although not statistically significant, interstitial inflammation was less intense in treated rats than non-treated controls. Glomerular expression of MMP-9 by immunoflourescence was significantly inhibited in the treated group. In addition pro-MMP-2 on gelatin zymography was importantly suppressed by doxycycline in ICN. CONCLUSION: Doxycycline, in addition to its antibiotic property, may, following further investigation, provide a possible survival benefit in proliferative glomerulonephritis.

In vitro reoxygenation following hypoxia increases MMP-2 and TIMP-2 secretion by human umbilical vein endothelial cells.

Endothelial cells lining the inner blood vessel walls play a key role in the response to hypoxia, which is frequently encountered in clinical conditions such as myocardial infarction, renal ischemia and cerebral ischemia. In the present study we investigated the effects of hypoxia and hypoxia/reoxygenation on gelatinases (matrix metalloproteinase-2 and -9), their inhibitor (TIMP-2) and activator (MT1-MMP), in human umbilical vein endothelial (HUVE) cells. HUVE cells were subjected to 4 h of hypoxia or hypoxia followed by 4 and 24 h of reoxygenation. The pro- and active forms of MMP-2 and MMP-9 were analyzed by gelatin zymography; TIMP-2 protein level was assayed using ELISA, while MT1-MMP activity was measured using an activity assay. The secretion of MMP-2 proform increased significantly in cells subjected to 4 h of hypoxia followed by 4 or 24 h of reoxygenation, compared with the normoxic group. TIMP-2 protein level also increased significantly in the hypoxia/reoxygenation groups, compared with the normoxic group. There were no statistically significant differences in the levels of active MT1-MMP in all groups. This study indicates that MMP-2 and TIMP-2 could be regarded as important components of a mechanism in the pathophysiology of ischemic injury following reperfusion.

MMP-2 and MMP-9 alteration in response to collaring in rabbits: the effects of endothelin receptor antagonism.

Matrix metalloproteinases (MMPs), and, in particular, gelatinases (MMP-2 and MMP-9), have been implicated in vascular cell proliferation and/or migration, contributing to intimal thickening, an essential stage in the development of atherosclerosis and restenosis following balloon angioplasty. Endothelin, a strong chemoatractant and mitogen, has been shown to promote smooth muscle cell proliferation and migration by activating MMPs via endothelin-A (ETA) receptors. The positioning of a soft silicon collar around the left carotid artery in rabbits results in intimal thickening. In this study, we investigate the possible role of gelatinases and the effect of a nonselective ETA/ETB receptor antagonist, TAK-044 (5 mg/kg body weight/day, subcutaneously [sc]), on these enzymes. Our results demonstrated that both MMP-2 and MMP-9 activities increased in response to collaring in placebo group, while treatment with TAK-044 significantly suppressed both gelatinase activities and proMMP-2 levels, and inhibited intimal thickening in collared arteries. These results suggest that either enhanced MMP expression or endothelin receptor antagonism may be involved in the formation of intimal thickening in this model.