Lysosomal Acid Phosphatase Activity Research Articles

Biochemical responses of the Dreissena polymorpha from municipal pond to caffeine, microplastics, and heating in single and combined exposures

Background. Pharmaceuticals have become the aquatic pollutants of growing concern. Caffeine is one of the most widely distributed in the surface waters among them. However, the environmentally relevant models of its effect, which include combined exposures with probable confounding factors, are unknown. Microplastics are a suspected vector that influences caffeine bioavailability. The temperature dependence of response, considering the increase of temperature in surface waters, can also be anticipated. The aim of this study was to analyze the input of caffeine, microplastics and elevated temperature into their combined effect on the zebra mussel Dreissena polymorpha (Pallas, 1771). Materials and Methods. Molluscs were exposed to caffeine (Caf, 20.0 μg·L−1), microplastics (MP, 1 mg·L−1, 2 μm in size), or elevated temperature (T, 25 °C) in the single and combined (Mix- and MixT-) exposures for 14 days. The concentrations of metallothioneins, metallothioneins-bound zinc, total Zn and Cu concentration in the tissue, total glutathione level, antioxidant (superoxide dismutase) and metabolic (citrate synthase) enzymes activities, acid phosphatase activity as the lysosomal functionality marker were determined. Results and Discussion. The decrease in Zn/Cu concentrations ratio in the soft tissues shared the common response in all exposures, reflecting the metal imbalance as the most sensitive marker. The MP-group was distinguished by the decrease in the levels of total Zn and extra lysosomal acid phosphatase activity, proving injury of Zn transportation and Zn-related enzyme activities. All other exposures (T-, Mix-, MixT-) caused citrate synthase and superoxide dismutase activation. Caf-related groups demonstrated the elevation of the levels of phosphatase lysosomal membrane-linked latency, metallothionein total protein and its apo-form. However, glutathione level was stable in all exposures. Conclusion. These data revealed the adverse effect of MP and shared beneficial effects in the exposures that involved caffeine, which can be explained by the antioxidant activity of this substance. Exposure to elevated temperature partially alleviated the effect of caffeine in the mixture. Thus, the results indicate the importance of multi-stress exposures modeling, which allows the evaluation of environmentally realistic responces of an organism to xenobiotics.

A new autophagy-driven protective pathway of Haematococcus astaxanthin against chemical damage in wistar rats with gentamicin-induced acute kidney injury

Astaxanthin was established to conserve kidney function and subcellular structure through anti-oxidation and/or the free radical scavenging system, yet little research linked a new protective effect to autophagy or lysosomes. We pre-fed Wistar rats with natural astaxanthin, β-carotene, or placebo and induced acute kidney injury using gentamicin, before examining renal tissues and measuring physiological indices. Qualitative evidence from histopathological and subcellular images, along with quantitative evidence showing treatment effects on blood urea nitrogen and serum creatinine (p < 0.01), indicated that esterified Haematococcus astaxanthin surpassed β-carotene at effectively counteracting chemical damage and protecting the kidneys from injury. Proliferation of enlarged lysosomes and mediation analysis results revealing enhanced lysosomal acid phosphatase activity were consistent with the hypothesized autophagy-lysosomal pathway being up-regulated by astaxanthin intake (p < 0.05). In conclusion, the protective effect of astaxanthin against acute kidney injury exerted through the autophagy-lysosomal detoxification pathway, which totally different from the anti-oxidation and/or conventional SOD-dependent free radical scavenging system, was demonstrated with strong evidence. In light of the pandemic outbreak of novel coronavirus pneumonia associated with a virus preferentially targeting the renal tubular cells, dietary astaxanthin may help bring down incidence rate of coronavirus disease, cases of acute kidney injury secondary to the disease, and mortality rate from acute kidney injury, especially when a standard of care treatment for the infectious disease is pending.

Are bio-based and biodegradable microplastics impacting for blue mussel (Mytilus edulis)?

The substitution of petrochemical plastics by bio-based and biodegradable plastics are in need of an evaluation for the potential toxic impacts that they can have on marine wildlife. This study aims to assess the toxicological effects of polylactic acid microparticles at two concentrations, 10 and 100 μg/L, during 8 days on the blue mussel, Mytilus edulis. No significant oxidative stress (catalase, glutathione-S-transferase and superoxide dismutase activities), neurotoxicity (acetylcholinesterase), or immunotoxicity (lysosomal membrane stability and acid phosphatase activity) were detectable. The multivariate analysis of metabolomic data allowed us to differentiate the individuals according to the exposure. From the loading plot of OPLS-DA, 48 ions down-regulated in the individuals exposed to microplastics. They were identified based on HRMS data as glycerophospholipids.

Comparison of markers of skin inflammation after injections of polylactic acid and threads based on polylactic acid

Aging processes lead to the tremendous changes in every tissues of human and skin is not an exception. Externally observable aging changes of the skin are caused by involutionary processes, which are happening on all levels, from skin to bones. The development of methods of prevention and treatment, which are aimed at fighting age-related changes in the skin and activating regenerative processes, is a live issue and has great scientific and practical significance.One of the important factors of skin aging is involutional changes occurring in the dermis. Currently, injections, based on polylactic acid are actively used as collagen stimulants in aesthetic medicine. Polylactic acid is a hydrophobic, biodegradable polymer derived from lactic acid. However, the mechanism of effect of polylactic acid on the components of the dermis, as well as the morphohistochemical status of the implant itself and the effect on markers of skin inflammation remains insufficiently understood and are actively discussed in the literature.We carried out biochemical studies of rat skin in order to study the effect of polylactic acid on markers of skin inflammation. To assess the condition of the skin of rats, after the administration of the studied drugs, the following was determined:• marker of inflammation - the activity of neutrophilic elastase;• factor of the state of cell membranes - the activity of lysosomal acid phosphatase;• the state of nonspecific antimicrobial protection - the activity of lysozyme.The conducted study allows us to conclude about the relative safety of preparations based on polylactic acid, the subdermal administration of which after 60 days shows minor signs of inflammation and practically does not affect the nonspecific antimicrobial protection of the skin of laboratory rats.

Pantoprazole pretreatment elevates sensitivity to vincristine in drug-resistant oral epidermoid carcinoma in vitro and in vivo

Resistance to chemotherapeutic agents is a major cause of treatment failure in patients with oral cancer. Proton pump inhibitors (PPIs), essentially H+-K+-ATPase inhibitors which are currently used in the treatment of acid related diseases, have demonstrated promising antitumor and chemo-sensitizing efficacy. The main purpose of the present study was to investigate whether pantoprazole (PPZ, one of PPIs) could increase the sensitivity of chemoresistant oral epidermoid carcinoma cells (KB/V) to vincristine (VCR) and elucidate the underlying action mechanism. Results showed that combination treatment of PPZ and VCR synergistically inhibited the proliferation of KB/V cells in vitro and in vivo. Furthermore, administration of PPZ and VCR not only induce apoptosis and G2/M phase arrest in KB/V cells but also suppress the migration and invasion of KB/V cells. The mechanism underlying synergistic anti-tumor effect of PPZ and VCR was related to the inhibition of the function and expression of P-glycoprotein (P-gp) and the down-regulation of EGFR/MAPK and PI3K/Akt/mTOR signaling pathways in KB/V cells. Additionally, we observed that PPZ treatment induced an increase in lysosomal pH and inhibited the activity of lysosomal enzyme acid phosphatase in KB/V cells, which could functionally reduce the sequestration of VCR in lysosomes and sensitized KB/V cells to VCR. In conclusion, our study demonstrated that PPZ could be included in new combined therapy of human oral cancer (especially on VCR-resistant therapy) together with VCR.

Phagolysosomal activity of macrophages in Nile tilapia (Oreochromis niloticus) infected in vitro by Aeromonas hydrophila: Infection and immunotherapy

The biochemical mechanisms involved in phagocytosis and the intracellular survival of Aeromonas hydrophila (Ah) in host macrophages (MΦs) are complex processes that affect infection success or failure. Thus, in the present study, we described the in vitro infection of Nile tilapia MΦs by a homologous bacterium and tested the effects of anti-A. hydrophila immunoglobulin Y (IgY) on the phagolysosomal activity and intracellular survival of the pathogen. The anti-Ah IgY modulated lysosomal acid phosphatase (LAP) activity as well as the production of reactive oxygen intermediates (ROIs) and nitric oxide (NO), thereby potentiating phagocytosis and the elimination of Ah. Thus, we assume that the specific IgY had a beneficial effect on infection control and postulated the use of the Nile tilapia MΦs as an important in vitro experimental model for the functional and therapeutic study of Ah infection.

Effect of L-arginine and carnitine on cathepsin L and H activity and lysosomal membranes permeability in myocardium in expressed hyperhomocysteinemia

Aim. To study the activity of lysosomal cysteine proteases (cathepsins L, H) and acid phosphatase, changing of permeability, stability of myocardial lysosomal membranes in rats in experimental expressed hyperhomocysteinemia model, and while administering L-arginine and carnitine. Methods. The study was performed on male Wistar rats kept on standard vivarium conditions divided into three control and three experimental groups of 8 animals each. Experimental samples were administered methionine, or combination of L-arginine and carnitine with methionine. The level of serum homocysteine was measured by ELISA. Cathepsin L and H activity was detected by spectrofluorimetric method. Acid phosphatase activity was recorded using the «end point» method. Results. In the model of expressed hyperhomocysteinemia the increase of cathepsin H total activity due to both lysosomal and nonlysosomal fractions was found. These changes were observed along with the general increase of lysosomal membranes permeability. When correcting hyperhomocysteinemia with L-arginine and carnitine a decrease of cathepsin L and H levels was noted as well as positive effect on the myocardial lysosomal membranes stability. Conclusion. Expressed hyperhomocysteinemia is accompanied by statistically significant increase of both lysosomal and cytoplasmic fractions of the cathepsin H activity, indicating the lysosomal membranes permeabilisation phenomenon; L-carnitine and arginine correct hyperhomocysteinaemia effects, leading to cathepsin L and H reduced activity and having a stabilizing effect on the lysosomal membranes of cardiomyocytes.

Cytotoxicity and cellular mechanisms involved in the toxicity of CdS quantum dots in hemocytes and gill cells of the mussel Mytilus galloprovincialis

CdS quantum dots (QDs) show a great promise for treatment and diagnosis of cancer and for targeted drug delivery, due to their size-tunable fluorescence and ease of functionalization for tissue targeting. In spite of their advantages it is important to determine if CdS QDs can exert toxicity on biological systems. In the present work, cytotoxicity of CdS QDs (5nm) at a wide range of concentrations (0.001–100mg Cd/L) was screened using neutral red (NR) and thiazolyl blue tetrazolium bromide (MTT) assays in isolated hemocytes and gill cells of mussels (Mytilus galloprovincialis). The mechanisms of action of CdS QDs were assessed at sublethal concentrations (0.31–5mg Cd/L) in the same cell types through a series of functional in vitro assays: production of reactive oxygen species (ROS), catalase (CAT) activity, DNA damage, lysosomal acid phosphatase (AcP) activity, multixenobiotic resistance (MXR) transport activity, Na-K-ATPase activity (only in gill cells) and phagocytic activity and damage to actin cytoskeleton (only in hemocytes). Exposures to CdS QDs lasted for 24h and were performed in parallel with exposures to bulk CdS and ionic Cd. Ionic Cd was the most toxic form to both cell types, followed by CdS QDs and bulk CdS. ROS production, DNA damage, AcP activity and MXR transport were significantly increased in both cell types exposed to the 3 forms of Cd. CAT activity increased in hemocytes exposed to the three forms of Cd while in gill cells only in those exposed to ionic Cd. No effects were found on hemocytes cytoskeleton integrity. Effects on phagocytosis were found in hemocytes exposed to bulk CdS and to CdS QDs at concentrations equal or higher than 1.25mg Cd/L but not in those exposed to ionic Cd, indicating a particle-specific effect on phagocytosis. In conclusion, cell-mediated immunity and gill cell function represent significant targets for CdS QDs toxicity.

ZnT2 accumulates zinc in lysosomes and mitochondria to signal mammary gland involution

Post‐lactation, the mammary gland (MG) undergoes massive apoptotic‐ and lysosomal‐mediated cell death and involutes back to the pre‐pregnant state. Recent data from our lab suggests zinc (Zn) import into specific intracellular compartments regulates this process. We found that the Zn transporter ZnT2 imports Zn into vesicles, which is required for Zn transfer into milk during lactation. In addition, ZnT2 imports Zn into mitochondria, which decreases ATP and activates apoptosis in mammary epithelial cells (MECs). Thus, we hypothesized that ZnT2‐mediated Zn accumulation into lysosomes and mitochondria signals MG involution (INVO). We determined that ZnT2 accumulates in lysosomes using sucrose gradient fractionation of MG from involuting mice. This increased lysosomal Zn and activity of the Zn‐dependent lysosomal enzyme acid phosphatase (AP), a marker of INVO. ZnT2 overexpressed in MECs in vitro co‐localized with lysosomes (LAMP1) and mitochondria (Mitotracker) and increased AP activity, apoptosis and cell death. ZnT2 overexpressed in MG using a recombinant adenovirus in vivo increased mitochondrial Zn, activated apoptosis and increased AP activity, which activated premature INVO. Collectively, these data suggest Zn accumulation into lysosomes and mitochondria acts as a regulatory signal to induce MG INVO. Dysregulation in this process may lead to premature INVO and suboptimal lactation.Grant Funding Source: Supported by NIH R01 HD058614 to SLK

Generation of free radicals in haemocytes of mussels after exposure to low molecular weight PAH components: Immune activation, oxidative and genotoxic effects

Polycyclic aromatic hydrocarbons (PAHs) constituents, such as phenanthrene (PH) and anthracene (AN), are considered toxic for marine organisms, including bivalve mollusks. The present study showed that the perturbation of health status in mussels, as well as their inability to survive in air (stress on stress response), following exposure to either PH or AN (at a final concentration of 0.1mg/L respectively), and a mixture of them (ration 1:1, at a final concentration of 0.2mg/L) for 7days, is probably related with alterations occurred in their haemocytes. According to the present study, PH and AN, either alone or in a mixture, could induce elevated levels of superoxide anions (•O2−) within haemocytes of mussels, thus leading to immune susceptibility as indicated by the increased levels of cell death and the elevated levels of lysosomal membrane acid phosphatase (AcP) activity, probably occurred via its overproduction or its release into the cytosol after lysosomal membrane destabilisation. PAH-mediated oxidative and genotoxic effects, indicated by the increased levels of both lipid peroxidation (MDA content) and nuclear abnormalities (MN test) could be related with haemocytes' inability to overcome free radical generation, thus leading to attenuation of general health status, before other disturbances, such as death, occur.

Aflatoxin B1 induced alterations in the stability of the lysosomal membrane in Oreochromis mossambicus (Peters, 1852)

This article elucidates the effect of crude dietary aflatoxin B1 (AFB1) on the stability of the lysosomal membrane in tilapia (Oreochromis mossambicus), and the analysis was made at the biochemical and cellular level. The liver lysosomal acid phosphatase activity was used to evaluate the functional integrity of the lysosome. Three experimental groups were fed on diets containing 0.375, 2.5 and 6 mg kg−1 of AFB1 for a time period of 6 weeks, whereas a fourth group fed on the semi-purified diet formed the control. Significant changes were observed in the stability of the lysosomal membrane in aflatoxin exposed groups. This is the first report, so far, of changes induced by AFB1 on the stability of lysosomal membrane in tilapia (O. mossambicus).

Dietary microorganism and plant effects on the survival and immune response of Litopenaeus vannamei challenged with the white spot syndrome virus

The effect of plants and probiotics on the survival and immune response of Litopenaeus vannamei challenged with the white spot syndrome virus (WSSV) was evaluated. A probiotic mixture (PM), plant extract (PE) or powdered plants (PP) were added to feed with the attractant Dry Oil®. An experiment was conducted with five treatments in triplicate. Shrimp (weighing 11.70±2.5 g) were cultured in 120 L plastic tanks and fed twice a day with commercial feed plus additives or with commercial feed plus WSSV. Animals were monitored for the occurrence of WSSV using single-step and nested PCR. The PM and PP added to the commercial feed showed high survival, a decrease in WSSV prevalence in shrimp and an increase in the activity of lysosomal enzymes, N-acetyl-β-glucosaminidase and acid phosphatase. The total haemocyte count in shrimp treated with PM was significantly higher than that in the control group (treatment I) and in shrimp fed with PE. The results of the present work indicate that PP and PM are good candidates for use as feed additives against WSSV in shrimp cultures.

Lysosomal acid phosphatase mediates dedifferentiation in the regenerating salamander limb

In this study, monoclonal antibodies against lysosomal acid phosphatase (LAP) of a salamander, Hynobius leechii, were used to determine the spatial and temporal expression of the LAP in the regenerating limbs. The Western blot and immunohistochemical analysis in the limb regeneration revealed that LAP was highly expressed at the dedifferentiation stage, especially in the wound epidermis and dedifferentiating limb tissues such as muscle and cartilage. With RA treatment, the LAP expression became upregulated in terms of both level and duration in the wound epidermis, blastemal cell and dedifferentiating limb tissues. In addition, in situ activity staining of LAP showed a similar result to that of immunohistochemistry. Thus, the activity profile of LAP activity coincides well with the expression profile of LAP during the dedifferentiation period. Furthermore, to examine the effects of lysosomal enzymes including LAP on salamander limb regeneration, lysosome extract was microinjected into limb regenerates. Interestingly, when the lysosome extract was microinjected into limb regenerates with a low dose of RA (50 µg/g body wt.), skeletal pattern duplication occurred frequently in the proximodistal and transverse axes. Therefore, lysosomal enzymes might cause the regenerative environment and RA plays dual roles in the modification of positional value as well as evocation of extensive dedifferentiation for pattern duplication. In conclusion, these results support the hypothesis that dedifferentiation is a crucial event in the process of limb regeneration and RA-evoked pattern duplication, and lysosomal enzymes may play important role(s) in this process.

Krüppel-like Factor 6 (KLF6) Affects the Promoter Activity of the α1-Proteinase Inhibitor Gene

Keratoconus is a progressive disease that thins and scars the cornea. In keratoconus corneas, levels of degradative enzymes, including lysosomal acid phosphatase (LAP) and cathepsin B, are elevated, and those of inhibitors alpha1-proteinase inhibitor (alpha1-PI) and alpha2-macroglobulin (alpha2-M) are reduced. The present study explored the possible involvement in keratoconus of Krüppel-like factor 6 (KLF6), a transcription factor previously described to be essential for the integrity of the corneal epithelium. The transcript and proteins level of KLF6 and its action in regulating the genes affected in keratoconus were examined in this study. Semiquantitative RT-PCR, Western blot analysis, immunofluorescence and in situ hybridization were used to investigate the expression of KLF6 mRNA and protein in normal and keratoconus corneas. Modulation by KLF6 of the promoter activity of alpha1-PI, LAP, cathepsin B, and alpha2-M genes was studied after transient transfection of KLF6 expression plasmid into corneal epithelial cells using promoter-reporter gene assays. Chromatin immunoprecipitation (ChIP) assays were performed to confirm the interactions between KLF6 and promoters of the genes affected in keratoconus. A global increased expression of the transcription factor KLF6 in terms of mRNAs and proteins was observed in total cornea and/or the epithelium in a substantial number of the keratoconus specimens. The promoter activity of the human alpha1-PI gene was suppressed by expression of KLF6 in corneal epithelial cells. The ChIP assay confirmed a physical interaction between KLF6 and the alpha1-PI promoter. Transcription factor KLF6 downregulates the alpha1-PI gene in corneal epithelial cells and may thereby be involved in keratoconus.

Procoagulant potential of platelet α granules

In this brief review of the literature it is pointed out that during platelet activation and degranulation platelet α granules leave the platelet interior through blebs in platelet plasma membrane and through the tips of the pseudopods, and then accumulate in the external milieu. There they undergo disintegration and secondary adhesion to the platelet plasma membranes. During their disintegration they expose their tightly packed GPIIb–IIIa complexes, annexin V stainable aminophospholipids, factor V, and the membrane markers CD62 and CD63. There is also demasking of lysosomal acid phosphatase activity and microvesicle formation. Lysosomal nature of platelet α granules is mentioned. It is suggested that platelet α granules are the sole source of platelet procoagulant activity and platelet microparticles (MP) or microvesicles (MV). The implications of this concept for antiplatelet therapy are discussed. A relationship of this process to tissue factor exposure and apoptosis is suggested.

Haemoglobin and Lung Total and Lysosomal Phosphatase Activity response to Passive Cigarette Smoking in the Albino Rat

Smoking of ordinary cigarette exposes its active smokers to health hazards and the filter-tipped cigarette, is said to reduce the hazards. However, it is not certain if the filter-tipped cigarette reduces the hazards in passive smokers since they inhail the cigarette smoke through their nostrils regardless of whether the cigarette is filter-tipped or not. Comparisons were made of the effect of Benson & Hedges cigarette (filter-tipped) and Captain Black cigar (Non filter-tipped) smoke on rat plasma haemoglobin, carboxy-haemoglobin and nicotine levels, average body weight and daily feed intake, lung Total and lysosomal acid phosphatase activity. The results showed that passive smoking reduced body weight and caused elevation of plasma haemoglobin, carboxyhaemoglobin and nicotine levels. Total lung acid phosphatase activity and lysosomal acid phosphatase activity. Filter-tipped Benson & Hedges cigarette smoke curtailed plasma carboxyhaemoglobn level, total lung acid phosphatase activity and lysosomal acid phosphatase activity and caused elevation of plasma nicotine level compared with non-filter-tipped Captain Black cigar smoke. KEY WORDS: Cigarette smoking, carboxyhaemoglobin; Total and lysosomal acid phosphatase activity. Nigerian Journal of Health and Biomedical Sciences Vol.3(2) 2004: 123-130

Green tea extract attenuates cyclosporine A-induced oxidative stress in rats

Cyclosporine A (CsA) nephrotoxicity underweighs the therapeutic benefits of such a powerful immunosuppressant. Whether oxidative stress plays a role in such toxicity is not well delineated. We investigated the potential of green tea extract (GTE) to attenuate CsA-induced renal dysfunction in rats. Three main groups of Sprague–Dawley rats were used: CsA, GTE, and GTE plus CsA-receiving animals. Corresponding control groups were also used. CsA was administered in a dose of 20 mg kg −1 day −1, i.p., for 21 days. In the GTE/CsA groups, the rats received different concentrations of GTE (0.5, 1.0 and 1.5%), as their sole source of drinking water, 4 days before and 21 days concurrently with CsA. The GTE group was treated with 1.5% concentration of GTE only for 25 days. A concomitant administration of GTE, to CsA receiving rats, markedly prevented the generation of thiobarbituric acid-reacting substances (TBARS) and significantly attenuated CsA-induced renal dysfunction as assessed by estimating serum creatinine, blood urea nitrogen, uric acid and urinary excretion of glucose. A considerable improvement in terms of reduced glutathione content and activity of antioxidant enzymes in the kidney homogenate of the GTE/CsA-receiving rats was observed. The activity of lysosomal enzymes, N-acetyl-β-glucosaminidase, β-glucuronidase and acid phosphatase was significantly inhibited following GTE co-administration. Our data prove the role of oxidative stress in the pathogenesis of CsA-induced kidney dysfunction. Supplementation of GTE could be useful in reducing CsA nephrotoxicity in rats. However, clinical studies are warranted to investigate such an effect in human subjects.

Tartrate-resistant acid phosphatase knockout mice.

TRACP is a lysosomal enzyme found in diverse tissues, where it is expressed in dendritic cells as well as osteoclasts and macrophages. To investigate the function of TRACP in vivo, we have generated mice in which the gene-encoding TRACP has been selectively disrupted by targeted homologous recombination in murine embryonic stem cells. Homozygous TRACP "knockout" mice have progressive foreshortening and deformity of the long bones and axial skeleton suggesting a role for TRACP in endochondral ossification. There is increased mineralization reflecting a mild osteopetrosis caused by reduced osteoclast modeling activity. These knockout mice also display an impairment of macrophage function with abnormal immunomodulatory cytokine responses. Superoxide formation and nitrite production were enhanced in stimulated macrophages lacking TRACP as was the secretion of the proinflammatory cytokines TNF-alpha, interleukin (IL)-1beta, and IL-12. TRACP knockout mice showed delayed clearance of the microbial pathogen Staphylococcus aureus after sublethal intraperitoneal inoculation. The macrophages lacking TRACP showed an increase in tartrate-sensitive lysosomal acid phosphatase activity (LAP). The TRACP knockout mice were bred with mice lacking LAP. Mice lacking both TRACP and LAP had even shorter bones than the TRACP single knockouts. Osteopontin, identical to the T-cell cytokine eta-1, accumulated adjacent to actively resorbing osteoclasts suggesting that both phosphatases are important for processing this protein. We propose that TRACP may be an important regulator of osteopontin/eta-1 activity common to both the immune system and skeleton.

Arrested apoptosis without nuclear fragmentation produced by efferent duct ligation in round spermatids and multinucleated giant cells of rat testis.

The apoptotic process evoked by efferent duct ligation in the testes of adult rats was followed for 10 days by differential staining for haematoxylin-eosin, periodic acid-Schiff and a modified trichrome technique in optical microscopy and by ultrastructural localization of acid phosphatase. Round spermatids showed the first effects of efferent duct ligation. At day 3 after ligation, annular clumps of chromatin with typical apoptotic characteristics appeared against the nuclear membrane of these cells. Afterwards, membranous structures and a wide separation between the two layers of the nuclear membrane were observed but nuclear fragmentation did not occur and apoptotic granules were not seen. Cytoplasmic components were also altered, and severely damaged organoids and empty vacuoles lacking acid phosphatase reaction were frequently seen. On day 2 after efferent duct ligation, multinucleated giant cells appeared, which displayed similar characteristics as spermatids and showed no acid phosphatase reaction. Although abnormal spermatids and multinucleated giant cells were surrounded by the cytoplasm of Sertoli cells, neither lysosomal acid phosphatase nor phagocytic activity was detected. It is concluded that efferent duct ligation specifically affects round immature spermatids eliciting a partial nuclear apoptotic response that is not accompanied by autophagic or heterophagic activity and without lysosomal participation in Sertoli cells.

Inhibition of human renal acid phosphatases by nephrotoxic micromolar concentrations of fluoride

Fluoride is considered to be a nephrotoxic substance, due to the association of 50-180 microM serum concentrations of fluoride with dose-related, subclinical to overt clinical renal impairment. At these concentrations the cellular targets of fluoride in renal tissue remain unknown. Fluoride at micromolar concentrations inhibits some enzymes including phosphatases. Here the effects of fluoride on a recently characterized acid phosphatase complex present only in a few human tissues were studied. This enzyme complex consists of alkaline fixation-resistant beta-glycerophosphatase and tartrate-resistant a-naphthyl phosphatase, and these are different from activities of known types of acid phosphatase and specific phosphatases. In kidney, strong activities for this complex are detected only in the endothelium of the afferent arterioles and in glomeruli. It appeared that alkaline fixation-resistant and lysosomal acid phosphatase activities were significantly inhibited in afferent arterioles and glomeruli by 75 microM fluoride. Tartrate-resistant activity was significantly inhibited by greater concentrations (250 microM). Inhibition of acid phosphatases in the afferent arterioles and glomeruli may be one renal target of fluoride nephrotoxicity. Although the physiological substrates of this acid phosphatase complex are unknown, its specific and restricted location may indicate a role in regulation of renal circulation.