ALLOSTERIC MODULATION BY ATP, CALCIUM AND MAGNESIUM-IONS OF RAT OSSEOUS PLATE ALKALINE-PHOSPHATASE

Alkaline phosphatase from rat osseous plate is allosterically modulated by ATP, calcium and magnesium at pH 7.5. At pH 9.4, the hydrolysis of ATP and PNPP follows Michaelis-Menten kinetics with K0.5 values of 154 muM and 42 muM, respectively. However, at pH 7.5 both substrates exhibit more complex saturation curves, while only ATP exhibited site-site interactions. Ca2+-ATP and Mg2+-ATP were effective substrates for the enzyme, while the specific activity of the enzyme for the hydrolysis of ATP at pH 7.5 was 800-900 U/mg and was independent of the ion species. ATP, but not PNPP, was hydrolyzed slowly in the absence of metal ions with a specific activity of 140 U/mg. These data demonstrate that in vitro and at pH 7.5 rat osseous plate alkaline phosphatase is an active calcium or magnesium-activated ATPase.

THE PHO-2A MUTANT OF NEUROSPORA-CRASSA WHICH IS DEFICIENT IN PI-REPRESSIBLE ALKALINE-PHOSPHATASE (EC 3.1.3.1) IS ALSO DEFECTIVE IN PI-REPRESSIBLE ACID-PHOSPHATASE (EC 3.1.3.2)

1. The mycelial Pi-repressible acid phosphatase presented p-nitrophenylphosphatase activity with negative cooperativity and Michaelian behavior when synthesized by the wild-type and pho-2A mutant strains of Neurospora crassa, respectively.2. The major acid phosphatase present in cell extracts of the pho-2A mutant of N. crassa grown in low Pi medium is more thermolabile (t1/2 = 4 min at 54-degrees-C, pH 5.4) than that of the wild strain (stable for at least 80 min at 54-degrees-C, pH 5.4).3. The pho-2A mutant of N. crassa secreted a more thermolabile acid phosphatase (t1/2 = 30 min at 50-degrees-C, pH 5.4) than the wild strain (t1/2 of at least 80 min at 50-degrees-C, pH 5.4).4. The pho-2A mutant of N. crassa synthesized a more thermolabile acid phosphatase (t1/2 = 37 min at 54-degrees-C, pH 5.4) than the wild strain in high Pi medium (t1/2 = 14 min al 54-degrees-C, pH 5.4).5. The pleiotropic nature of the pho-2 locus and its possible involvement in the mechanism of phosphatase secretion by N. crassa are proposed.

Effects of Chronic Physical Activity and of Ultrasound Treatment on Bone Consolidation

The objective of this study was to investigate the effects of ultrasound treatment and physical exercise on the velocity of bone consolidation and resistance to deformation. We performed osteotomy in the upper third of the right tibia of rats. Physical training consisted of swimming 1 h per day with a load of 5% b.w. for 30 days. Therapy with medium-intensity ultrasound was applied daily on the damaged area. Wistar rats were divided into the following groups: osteotomized sedentary animals with no ultrasound treatment (1.OSnUS), osteotomized trained animals with no ultrasound treatment (2.OTnUS), osteotomized sedentary animals with ultrasound treatment (3.OSwUS). and osteotomized trained animals with ultrasound treatment (4.OTwUS). The animals were sacrificed for the following analyses: muscle glycogen, serum alkaline phosphatase at the 5th, 10th, 20th, and 30th days, test of maximum resistance to flexion, rupture flexion and mean tibial rigidity at the 30th day. Muscle glycogen was increased at the 20th day: alkaline phosphatase was elevated at the 5th and 20th days in groups 3.OSwUS and 4.OTwUS. and decreased at the 10th day. Groups1.OSnUS and 2.OTnUS did not show significant variations. In the mechanical resistance tests, we noted that ultrasound therapy and the association of physical activity used in the present study showed significant differences in bone resistance and bone rigidity after 30 days of treatment. These facts suggest that ultrasound or physical activity, or their combination may accelerate the process of bone tissue repair.

Enzymatic activity of hypopharyngeal gland extracts from workers of Apis mellifera (Hymenoptera, Apidae, Apinae)

This research presents a comparative study of enzymatic activity of the hypopharyngeal gland extracts from workers of Apis mellifera in three physiologic stages: newly emerged, nurse and forager workers, with the objective of contributing to the comprehension of the gland function. In order to determinate the enzymes present in the extracts, the Api Zym kit (Bio Mérieux) was used to test the activity of 19 different enzymes. The enzymes found in larger amounts only in the hypopharyngeal glands from certain individuals were the following: in newly emerged workers, the N-acetyl-down double arrow sign-glucosaminidase that may be digesting the chitin of some food ingested by the bee; in forager workers, the acid phosphatase that is likely acting in authophagic processes, the a-glucosidase, in the processing of nectar into honey, and the down double arrow sign-glucosidases, in the pollen digestion.

A comparative study of protein and enzymatic activity in venoms of some common wasps (Hymenoptera: Vespidae) from São Paulo State

The Hymenoptera Aculeata venoms, with few exceptions, have been poorly studied and characterized. Nevertheless, they have raised increasing interest due to their medical importance, since accidents with these insects are fairly frequent in Brazil and may cause severe allergic reactions. The objectives of the present work were the quantitative characterization of the main allergenic enzymes present in the venom of the species Polybia paulista, Polybia ignobilis, Polistes simillimus, and Agelaia pallipes pallipes through biochemical assays for the determination of total protein content, as well as the level of the enzymatic activity of phospholipase, hyaluronidase, acid phosphatase and esterase. These results, in addition to providing biochemical knowledge about the venom of the species in question, also supply studies that allow phylogenetic inferences among them.

Rhipicephalus (Boophilus) microplus (Canestrini, 1887) (Acari: Ixodidae): Acid phosphatase and ATPase activities localization in salivary glands of females during the feeding period

This study investigates the presence and the localization of acid phosphatase and ATPase in the salivary glands of Rhipicephalus (Boophilus) microplus female ticks during feeding. Semi-engorged females showed a larger amount of acid phosphatase compared to those at beginning of feeding, localized mainly in the apical portion of the secretory cells, and in the basal labyrinth of the interstitial cells. Ultrastructural observations also demonstrated its presence in secretion granules and inside some nuclei of secretory cells at beginning of feeding. Acid phosphatase in a free form probably has a hemolymph and/or ribosomal origin and participates in salivary gland secretion control. ATPase was detected in basal membrane of all types of acini and/or in the cytoplasm of the secretory cells at both feeding stages. The enzyme activities found strongly suggests that cell death by apoptosis occurs during the degenerative process. © 2006 Elsevier Inc. All rights reserved.

Alkaline phosphatase localization in golden hamster epididymidis: A biochemical and histochemical investigation

The aim of this study was to determine alkaline phosphatase (ALP; E.C. 3.1.3.1) activity and major expression in homogenates obtained from different regions of Golden hamster epididymis, comprising the initial segment, head, body and tail, with concomitant research of this enzyme localization and activity in samples of tissues. These were collected from the same regions and investigated by histochemical conventional study performed on frozen histological sections. No significant differences in mean ALP activity, reported as U.100 mg-1 of tissue, were observed among the biological specimens collected from the epididymidis initial segment (0.92 ± 0.28 U.100 mg-1 tissue), head (1.07 ± 0.67 U.100 mg-1 tissue) and body (0.77 ± 0.23 U.100 mg-1 tissue). However, mean ALP activity was significantly higher in the epididymal tail (8.94 ± 0.40 U.100 mg-1 tissue) compared with the precedent segments. The findings suggested that ALP plays a significant role in the tail of the Golden hamster epididymidis, mediating androgenic segregation necessary to maintain the epithelial integrity. Furthermore, ALP acts on active transport of substances between the luminal fluid and spermatozoon membrane, and contrariwise. Thus, the high concentration of ALP in the epididymal tail helps to indicate the importance of this enzyme in the metabolism and maintenance of spermatozoa maturation and storage into the epididymidis luminal compartment, perhaps directly influencing the normal reproductive morphophysiology.

Intestinal anti-inflammatory activity of coumarin and 4-hydroxycoumarin in the trinitrobenzenesulphonic acid model of rat colitis

Coumarins represent an important class of phenolic compounds with multiple biological activities, including inhibition of lipidic peroxidation and neutrophil-dependent anion superoxide generation, anti-inflammatory and immunosuppressor actions. All of these proprieties are essential for that a drug may be used in the treatment of inflammatory bowel disease. The present study examined intestinal anti-inflammatory activity of coumarin and its derivative, the 4-hydroxycoumarin on experimental ulcerative colitis in rats. This was performed in two different experimental settings, i.e. when the colonic mucosa is intact or when the mucosa is in process of recovery after an initial insult. The results obtained revealed that the coumarin and 4-hydroxycoumarin, at doses of 5 and 25 mg/kg, significantly attenuated the colonic damage induced by trinitrobenzenesulphonic acid (TNBS) in both situations, as evidenced macroscopically, microscopically and biochemically. This effect was related to an improvement in the colonic oxidative status, since coumarin and 4-hydroxycoumarin prevented the glutathione depletion that occurred as a consequence of the colonic inflammation. © 2008 Pharmaceutical Society of Japan.

Expression of acid phosphatase in the seminiferous epithelium of vertebrates

Acid phosphatases (AcPs) are known to provide phosphate to tissues that have high energy requirements, especially during development, growth and maturation. During spermatogenesis AcP activity is manifested in heterophagous lysosomes of Sertoli cells. This phagocytic function appears to be hormone-independent. We examined the expression pattern of AcP during the reproductive period of four species belonging to different vertebrate groups: Tilapia rendalli (Teleostei, Cichlidae), Dendropsophus minutus (Amphibia, Anura), Meriones unguiculatus (Mammalia, Rodentia), and Oryctolagus cuniculus (Mammalia, Lagomorpha). To demonstrate AcP activity, cryosections were processed for enzyme histochemistry by a modification of the method of Gömöri. AcP activity was similar in the testes of these four species. Testes of T. rendalli, D. minutus and M. unguiculatus showed an intense reaction in the Sertoli cell region. AcP activity was detected in the testes of D. minutus and O. cuniculus in seminiferous epithelium regions, where cells are found in more advanced stages of development. The seminiferous epithelium of all four species exhibited AcP activity, mainly in the cytoplasm of either Sertoli cells or germ cells. These findings reinforce the importance of AcP activity during the spermatogenesis process in vertebrates. © FUNPEC-RP.

Inhibition of osteoblast activity by zoledronic acid

INTRODUCTION: Patients treated with nitrogen-containing bisphosphonates, such as zoledronic acid (ZA), have frequently shown oral bone exposure areas, termed osteonecrosis. In addition, these patients may also present low repair and regeneration potential, mainly after tooth extractions. These side-effects caused by bisphosphonates may be due to their inhibitory effects on oral mucosa and local bone cells. OBJECTIVE: To evaluate the effects of ZA on the mineralization capacity of cultured osteoblasts. MATERIALS AND METHODS: Human immortalized osteoblasts (SaOs-2) were grown in plain culture medium (Dulbecco's Modified Eagle Medium [DMEM] + 10% fetal bovine serum [FBS]) in wells of 24-well plates. After 48-hour incubation, the plain DMEM was replaced by a solution with ZA at 5 µM which was maintained in contact with cells for seven, 14 or 21 days. After these periods, cells were evaluated regarding alkaline phosphatase (ALP) activity and mineral nodule formation (alizarin red). Data were statistically analyzed by Mann-Whitney test, at 5% of significance level. RESULTS: ZA caused significant reduction on ALP activity and mineral nodules formation by cultured osteoblasts in all evaluated periods (p < 0.05). CONCLUSION: These data indicate that ZA causes inhibition on the osteogenic phenotype of cultured human osteoblasts, which, in turn, may reduce bone repair in patients subjected to ZA therapy.

Exercise training decreases mitogen-activated protein kinase phosphatase-3 expression and suppresses hepatic gluconeogenesis in obese mice

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Hymenaea stigonocarpa Mart. ex Hayne: A tropical medicinal plant with intestinal anti-inflammatory activity in TNBS model of intestinal inflammation in rats

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Identification and enzymatic characterization of acid phosphatase from Burkholderia gladioli

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Antitumor activity of irradiated riboflavin on human renal carcinoma cell line 786-O

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Activation of the low molecular weight protein tyrosine phosphatase in keratinocytes exposed to hyperosmotic stress

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)