Potential of Vigna radiata (L.) sprouts in the management of inflammation and arthritis in rats: Possible biochemical alterations.

Vigna radiata (Fabaceae) is an important pulse crop widespread throughout the tropics and warm temperature regions. In this study, we evaluated the in vitro anti-inflammatory and in vivo antiarthritic activity of Vigna radiata sprouts in rats. The in vitro anti-inflammatory activity was determined by membrane stabilization and protein denaturation method. Whereas, the antiarthritic activity of the ethanolic extract of the sprouts was evaluated by complete Freund’s adjuvant model with diclofenac sodium as the standard drug. Body weights, paw volume, biochemical parameters such as lipid peroxidation, total reduced glutathione, myeloperoxidase and lysosomal enzymes like cathepsin-D, N-acetyl β-D-glucosamindase and β-D-glucuronidase were estimated. Treatment with ethanolic extract of V. radiata exhibited significant membrane stabilization activity and protein denaturation activity, and significantly attenuated the biochemical changes induced by administration of complete Freund’s adjuvant. The findings of the present study suggest the possible role of Vigna radiata in the therapeutics of arthritis.

A propósito de un caso de gangliosidosis GM-2 tipo II: enfermedad de Sandhoff / A propos of a case of gangliosidosis GM. Type II. Sandhoff disease

Las gangliosidosis son un conjunto de enfermedades hereditarias de almacenamiento lisosómico, debidas a un acúmulo de gangliósidos, sobre todo en las neuronas. La causa es la disfunción de alguna de las enzimas lisosómicas de la ruta de degradación de los gangliósidos. Existen varias formas de gangliosidosis, como son la GM1 y GM2. Se presentó el caso de una paciente de 33 años de edad, que había sido diagnosticada anteriormente de esclerosis lateral amiotrófica. Por varios síntomas presentados se le realizan una serie de exámenes complementarios, los cuales arrojan como resultado una gangliosidosis GM-2 tipo II o enfermedad de Sandhoff.

Gangliosidosis are a group of hereditary diseases of lysosomal storage, due to an accumulation of gangliosides, especially in the neurons. The cause is the dysfunction of several lysosomal enzymes in the way of the gangliosides degradation. There are several forms of gangliosidesis, like GM1 and GM2. We present the case of a 33-years-old patient who was previously diagnosed with lateral amyotrophic sclerosis. Because of several symptoms he presented we carried out some complementary exams showing as a result a gangliosidosis GM-2 Type II or Sandhoff disease.

Streptozotocin-induced diabetes mellitus affects lysosomal enzymes in rat liver

It has been previously shown that dextran sulfate administered to diabetic rats accumulates in the liver and kidney, and this could be due to a malfunction of the lysosomal digestive pathway. The aim of the present study was to evaluate the expression and activities of lysosomal enzymes that act upon proteins and sulfated polysaccharides in the livers of diabetic rats. Diabetes mellitus was induced by streptozotocin in 26 male Wistar rats (12 weeks old), while 26 age-matched controls received only vehicle. The livers were removed on either the 10th or the 30th day of the disease, weighed, and used to evaluate the activity, expression, and localization of lysosomal enzymes. A 50-60% decrease in the specific activities of cysteine proteases, especially cathepsin B, was observed in streptozotocin-induced diabetes mellitus. Expression (mRNA) of cathepsins B and L was also decreased on the 10th, but not on the 30th day. Sulfatase decreased 30% on the 30th day, while glycosidases did not vary (or presented a transitory and slight decrease). There were no apparent changes in liver morphology, and immunohistochemistry revealed the presence of cathepsin B in hepatocyte granules. The decrease in sulfatase could be responsible for the dextran sulfate build-up in the diabetic liver, since the action of sulfatase precedes glycosidases in the digestive pathway of sulfated polysaccharides. Our findings suggest that the decreased activities of cathepsins resulted from decreased expression of their genes, and not from general lysosomal failure, because the levels of glycosidases were normal in the diabetic liver.

Effect of Saccharum spontaneum Linn. on Lysosomal enzymes of Uro-lithiatic rats.

The ethanolic root extract of Saccharum spontaneum of family Poaceae was used to treatthe urolithiasis induced by glycolic acid On this course, the extract also repairs the changesthat happened in the lysosomal enzymes like β-D-glucuronidase, xanthine oxidase in liver and kidney and n-acetyl _-d-glucosaminidase in serum, liver, kidney and urine of the urolithiatic rats. The ethanolic root extract (200 and 300 / kg b.w.) elevated the levels of reduced β-D-glucuronidasein liver and n-acetyl _-d-glucosaminidase in liver and kidney and reduced the level of xanthineoxidase in liver and kidney and n-acetyl _-d-glucosaminidase in serum and urine significantly (p<0.05) when compared with the toxic groups. The results shown by the ethanolic root extract (200 and 300 mg / kg b.w.) was compared to standard thiazide drug treated group, showing no significantdifference (p<0.05) and thus it proves that the ethanolic root extract of S.spontaneum exhibits potent antiurolithiatic activity.

Efficacy of boswellic acid on lysosomal acid hydrolases, lipid peroxidation and anti-oxidant status in gouty arthritic mice

Objective:To evaluate the efficacy of boswellic acid against monosodium urate crystal-induced inflammation in mice. Methods:The mice were divided into four experimental groups. Group I served as control;mice in group II were injected with monosodium urate crystal;group III consisted of monosodium urate crystal-induced mice who were treated with boswellic acid (30 mg/kg/b.w.);group IV comprised monosodium urate crystal-induced mice who were treated with indomethacin (3 mg/kg/b.w.). Paw volume and levels/activities of lysosomal enzymes, lipid peroxidation, anti-oxidant status and inflammatory mediator TNF-αwere determined in control and monosodium urate crystal-induced mice. In addition, the levels of β-glucuronidase and lactate dehydrogenase were also measured in monosodium urate crystal-incubated polymorphonuclear leucocytes (PMNL) in vitro. Results:The activities of lysosomal enzymes, lipid peroxidation, and tumour necrosis factor-αlevels and paw volume were increased significantly in monosodium urate crystal-induced mice, whereas the activities of antioxidant status were in turn decreased. However, these changes were modulated to near normal levels upon boswellic acid administration. In vitro, boswellic acid reduced the level of β-glucuronidase and lactate dehydrogenase in monosodium urate crystal-incubated PMNL in concentration dependent manner when compared with control cells. Conclusions: The results obtained in this study further strengthen the anti-inflammatory/antiarthritic effect of boswellic acid, which was already well established by several investigators.

In vitro studies on inhibitory effect of proanthocyanidins in modulation of neutrophils and macrophages.

The role of proanthocyanidins (PC), a novel flavonoid extracted from grape seeds was studied in vitro in the modulation of neutrophil and macrophage function. We attempted to assess the levels of non-enzymatic and enzymatic mediators in the presence or absence of PC in 4-phorbol-12--myristate-13-acetate (PMA)-stimulated neutrophils isolated from humans and rats, E. coli endotoxin-stimulated macrophages and macrophages isolated from E. coli endotoxin-induced experimental periodontitis in rats. Addition of PC at a concentration of 50 µg/ml effectively blocked the release of reactive oxygen species (ROS) and reactive nitrogen species (RNS) and exhibited a marked inhibition of myeloperoxidase (MPO) and lysosomal enzymes (p<0.001), as compared to PMA-stimulated neutrophils (human and rats) and neutrophils isolated from experimental periodontitis in rats. The levels of ROS, RNS and lysosomal enzymes were found to be elevated (p<0.001) and addition of PC significantly (p<0.001) reduced these levels as compared to those from E. coli endotoxin-stimulatedmacrophages from rats and macrophages isolated from experimental periodontitis in rats (p<0.001). Thus, the study demonstrated that PC decreased the levels of ROS and RNS and also inhibited the MPO and lysosomal enzymes activities in experimental periodontitis in rats. In addition, this study clearly indicated that PC could be developed as an effective antiinflammatory agent.

Protective effect of proanthocyanidins on endotoxin induced experimental periodontitis in rats.

The pathogenesis of periodontitis involves anaerobic oral bacteria as well as the host response to infection and several drugs have been developed which can curtail these deleterious effects. Proanthocyanidin, a novel flavanoid extracted from grape seeds, has been shown to provide a significant therapeutic effect on endotoxin (Escherichia coli) induced experimental periodontitis in rats. In this study, protective action of different doses of proanthocyanidins was investigated in blood by assaying the reactive oxygen species such as hydrogen peroxide, superoxide anion, myeloperoxidase and lipid peroxides, lysosomal enzyme activities such as cathepsin B, cathepsin D, β-glucuronidase and acid phosphatase, nonenzymatic antioxidants such as ascorbic acid, -tocopherol, ceruloplasmin, reduced glutathione and antioxidant enzymes such as catalase, superoxide dismutase, glutathione peroxidase and glutathione-s-transferase. Experimental periodontitis rats showed a reduction in body weight and body weight gain could be noticed when they were administered proanthocyanidins. The levels of reactive oxygen species and lysosomal enzymes were found to increase whereas antioxidant levels were decreased significantly in experimental periodontitis. Proanthocyanidins at an effective dose of 30mg / kg body weight, sc, for 30 days effected a decrease in serum reactive oxygen species, lipid peroxides, lysosomal enzymes, acute phase proteins and an increase in antioxidant levels. Histopathological evidence of experimental periodontitis showed cellular infiltration of inflammatory cells while proanthocyanidin treated groups demonstrated only scattered inflammatory cells and blood vessels. Thus, the results showed that dietary supplementation of proanthocyanidin enhanced the host resistance as well as the inhibition of the biological and mechanical irritants involved in the onset of gingivitis and the progression of periodontal disease.

Mucopolysaccharidosis and mucolipidosis / 한양의대학술지

Mucopolysaccharidosis (MPS) and mucolipidosis(ML) belong to a group of rare genetic disorders of lysosomal enzymes and share some clinical manifestations. MPS is characterized by the accumulation of glycosaminoglycans (GAG) and results from the impaired function of one of 11 enzymes required for normal GAG degradation. ML, which is clinically similar to several forms of MPS, is caused by deficiency of Nacetylglucosamine-1-phosphotransferase activity. Therapeutic strategies for MPS, including enzyme replacement therapy and bone marrow transplantation, have been developed with some success. In this review, we discuss clinical feature, diagnostic methods, management and the present status of research on MPS and ML.

Plasma Activity of Lysosomal Enzymes in Active Pulmonary Tuberculosis / 결핵

BACKGROUND: The confirmative diagnosis of pulmonary tuberculosis(Tb) can be made by the isolation of Mycobacterium Tuberculosis(MTb) in the culture of the sputum, respiratory secretions or tissues of the patients, but positive result could not always be obtained in pulmonary Tb cases. Although there are many indirect ways of the diagnosis of Tb, clinicians still experience the difficulty in the diagnosis of Tb because each method has its own limitation. Therefore development of a new diagnostic tool is clinically urgent. It was reported that silica cause some lysosomal enzymes to be released from macrophages in vitro and one of these enzymes is elevated in workers exposed to silica dust and in silicotic subjects. In pulmonary Tb, alveolar macrophages are known to be activated after ingestion of MTb. Activated macrophages can kill MTb through oxygen free radical species and digestive enzymes of lysosome. But if macrophages allow the bacilli to grow intracellularly, the macrophages will die finally and local lesion will enlarge. Then it is assumed that the lysosomal enzymes would be released from the dead macrophages. The goal of this investigation was to determine if there are differences in the plasma activities of lysosomal enzymes, beta-glucuronidase(GLU) and beta-N- acetyl glucosaminidase(NAG), among the groups of active and inactive pulmonary Tb and healthy control, and to see if there is any possibility that the plasma activity of GLU and NAG can be used as diagnostic indicies of active pulmonary Tb. METHODS: The plasma were obtained from 20 patients with bacteriologically proven active pulmonary Tb, 15 persons with inactive Tb and 20 normal controls. In 10 patients with active pulmonary Tb, serial samples after 2 months of anti-Tb medications were obtained. Plasma GLU and NAG activities were measured by the fluorometric methods using 4-methylumbelliferyl sub- strates. All data are expressed as the mean +/-the standard error of the mean. RESULTS: The activites of GLU and NAG in plasma of the patients with active Tb were 21.52 +/-3.01 and 325.4+/-23.37(nmol product/h/ ml of plasma), respectively. Those of inactive pulmonary Tb were 24.87+/-3.78, 362.36+/-33.92 and those of healthy control were 25.45 +/-4.05, 324.44+/-28.66 (nmol product/ h/ml of plasma), respectively. There were no significant differences in the plasma activities of both enzymes among 3 groups. The plasma activities of GLU at 2 months after anti-Tb medications were increased(42.18+/-5.94 nmol product/h/ ml of plasma) in the patients with active pulmonary Tb compared with that at the diagnosis of Tb(P-value <0.05). CONCLUSION: The results of the present investigation suggest that the measurement of the plasma activities of GLU and NAG in the patients with active pulmonary Tb could not be a useful method for the diagnosis of active Tb. Further investigation is necessary to define the reasons why the plasma activities of the GLU was increased in the patients with active pulmonary Tb after Tb therapy.

Biochemical lesions in liver and kidney after caecal amoebiasis in rats by Entamoeba histolytica and their reversal by antiamoebic drugs.

Levels of lipid peroxides in rat caecum, blood, liver and kidney and the capacity of tissue homogenates to form lipid peroxides in vitro was enhanced after caecal amoebiasis in rats produced by Entamoeba histolytica (IB-1). The activity of hepatic drug-metabolizing enzymes in post-mitochondrial fraction and the cytochrome P 450 contents in microsomal fraction decreased significantly, while lysosomal enzymes such as acid phosphatase, acid ribonuclease and cathepsin Β showed an increase in the liver homogenates of infected animals. These changes were reversed following treatment with the antiamoebic drug, metronidazole.

Effects of protein deficiency on selective elicitation of lysosomal enzymes in rat peritoneal macrophages.

Young albino rats were fed ad libitum 4, 8 or 20 % (control) protein diet for 1–4 weeks. Total activities of some of the lysosomal enzymes, namely, acid phosphatase, aryl sulphatase, ß-glucuronidase and cathepsin D, were determined in resident and proteasepeptone elicited peritoneal macrophages. Total cell number, protein content and the lysosomal enzyme activities were increased significantly in protease-peptone elicited macrophages; though at a lower rate in 4 % protein-fed group compared to control ones. However, the rate of induction of the tested hydrolases was selective and their response to the stimulant varied widely. Similarly, response of each enzyme to low protein diet also varied. Thus, at 4 weeks, cathepsin D and ß-glucuronidase activities, expressed per total number of elicited macrophages were reduced by 45 and 60 %, respectively, in 4 % protein-fed animals. These results indicate that the metabolic events related to lysosomal function in macrophages, are affected by dietary restriction of proteins.

Studies of the synthesis, structure and function of the phosphorylated oligosaccharides of lysosomal enzymes.

Newly synthesized lysosomal enzymes were found to contain N-acetylglucosamine residues in phosphodiester linkage to the 6 position of the mannose residues on high-mannose type oligosaccharides. The formation of these structures was shown to be catalyzed by a specific N-acetylglucosaminylphosphotransferase enzyme, that utilises UDP-N-acetylglucosamine as a donor. The phosphorylation reaction can take place on any of four or five positions on the high-mannose oligosaccharide. Subsequently an α-N-acetylglucosaminylphosphodiesterase removes the outer blocking Nacetylglucosamine residues to generate the mature phosphomannsoyl recognition signal. This signal is responsible for the targetting of newly synthesized lysosomal enzymes to lysosomes. The human syndromes of I-cell disease (Mucolipidosis II) and pseudo-Hurler polydystrophy (Mucolipidosis III) were shown to be caused by deficiency of the first enzyme in the pathway, the UDP-N-acetylglucosamine: Glycoprotein N-acetylglucosaminylphosphotransferase.

Effect of cholesterol on multiplication, lipid metabolism and lysosomal enzymes of Hartmanella culbertsoni.

Addition of sonicated dispersions of cholesterol to peptone-salt-vitamin medium resulted in the metabolism of the sterol by Hartmanella culbertsoni. Trophozoite multiplication was stimulated at 1–5 mg/litre, but retarded at 10-20 mg/litre. When cholesterol was added to the medium, incorporation of [1,2—14C] –acetate into neutral lipid, phospholipid, non-saponifiable and cholesterol fractions of the amoebae was significantly reduced. Cholesterol ester was detected in the medium but phospholipids were not released. Addition of cholesterol stimulated the activity of lysosomal acid phosphatase, acid deoxyribonuclease and cathepsin Β but did not affect 5'-nucleotidase, adenosine triphosphatase, alkaline phosphatase, glucose-6-phosphatase, succinate dehydrogenase and cytochrome C oxidase.