Enhanced T-type calcium channel 3.2 activity in sensory neurons contributes to neuropathic-like pain of monosodium iodoacetate-induced knee osteoarthritis.

The monosodium iodoacetate knee osteoarthritis model has been widely used for the evaluation of osteoarthritis pain, but the pathogenesis of associated chronic pain is not fully understood. The T-type calcium channel 3.2 (CaV3.2) is abundantly expressed in the primary sensory neurons, in which it regulates neuronal excitability at both the somata and peripheral terminals and facilitates spontaneous neurotransmitter release at the spinal terminals. In this study, we investigated the involvement of primary sensory neuron-CaV3.2 activation in monosodium iodoacetate osteoarthritis pain. Knee joint osteoarthritis pain was induced by intra-articular injection of monosodium iodoacetate (2 mg) in rats, and sensory behavior was evaluated for 35 days. At that time, knee joint structural histology, primary sensory neuron injury, and inflammatory gliosis in lumbar dorsal root ganglia, and spinal dorsal horn were examined. Primary sensory neuron-T-type calcium channel current by patch-clamp recording and CaV3.2 expression by immunohistochemistry and immunoblots were determined. In a subset of animals, pain relief by CaV3.2 inhibition after delivery of CaV3.2 inhibitor TTA-P2 into sciatic nerve was investigated. Knee injection of monosodium iodoacetate resulted in osteoarthritis histopathology, weight-bearing asymmetry, sensory hypersensitivity of the ipsilateral hindpaw, and inflammatory gliosis in the ipsilateral dorsal root ganglia, sciatic nerve, and spinal dorsal horn. Neuronal injury marker ATF-3 was extensively upregulated in primary sensory neurons, suggesting that neuronal damage was beyond merely knee-innervating primary sensory neurons. T-type current in dissociated primary sensory neurons from lumbar dorsal root ganglia of monosodium iodoacetate rats was significantly increased, and CaV3.2 protein levels in the dorsal root ganglia and spinal dorsal horn ipsilateral to monosodium iodoacetate by immunoblots were significantly increased, compared to controls. Perineural application of TTA-P2 into the ipsilateral sciatic nerve alleviated mechanical hypersensitivity and weight-bearing asymmetry in monosodium iodoacetate osteoarthritis rats. Overall, our findings demonstrate an elevated CaV3.2 expression and enhanced function of primary sensory neuron-T channels in the monosodium iodoacetate osteoarthritis pain. Further study is needed to delineate the importance of dysfunctional primary sensory neuron-CaV3.2 in osteoarthritis pain.

Psoralen Protects Chondrocytes, Exhibits Anti-Inflammatory Effects on Synoviocytes, and Attenuates Monosodium Iodoacetate-Induced Osteoarthritis.

Current study examined whether psoralen (PSO) exhibits anti-inflammatory responses, protection and activation of chondrocytes, and relieve osteoarthritis (OA). Rats chondrocytes and human synoviocytes were cultured in tumor necrosis factor-α (TNF-α) conditioned culture medium with/without PSO to test the cell morphologies and cytotoxicities in vitro. Cartilaginous extracellular matrix (ECM) and proliferative gene/protein expression levels were evaluated in chondrocytes. Meanwhile, matrix metalloproteinases (MMPs) and interleukins (ILs) gene/protein expression were analyzed in synoviocytes. SD rats of monosodium iodoacetate (MIA) induced OA model were used in order to assess the effects of PSO on attenuating degeneration of the articular cartilage in vivo. Results showed TNF-α conditioned culturing with/without PSO (1-100 µM) had no any toxicity on both the cell lines. PSO (10 µM) activated cartilaginous specific ECM expression along with up-regulation of proliferative genes at transcriptional levels. Interestingly, PSO significantly reversed TNF-α induced up-regulation of MMP13 and ILs synoviocytes in a dose-dependent manner (1 to 20 µM), while down-regulated cartilaginous ECM production. Following six weeks of PSO treatments to articular cartilage osteoarthritis, compared to MIA-induced group, the appearance and physiological structure of articular cartilage was more integrated with greatly organized chondrocytes and abundant cartilage matrix. In conclusion, PSO protects and activates chondrocytes, antagonizing the expression of MMPs and ILs secreted by synovial cells, and effectively attenuates MIA-induced OA.

Sodium Acetate, Sodium Acid Pyrophosphate, and Citric Acid Impacts on Isolated Peripheral Lymphocyte Viability, Proliferation, and DNA Damage.

The present study examined the impacts of sodium acetate (SA), sodium acid pyrophosphate (SAPP), and citric acid (CA) on the viability, proliferation, and DNA damage of isolated lymphocytes in vitro. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) release assays were adopted to evaluate cell viability, while comet assay was employed to assess the genotoxic effects. The cells were incubated with different levels of SA (50, 100, and 200 mM), SAPP (25, 50, and 100 mM/L), or CA (100, 200, and 300 µg/mL). The lymphocytes treated with the tested food additives showed concentration-dependent decreases in both cell viability and proliferation. A concentration-dependent increase in LDH release was also observed. The comet assay results indicated that SA, SAPP, and CA increased DNA damage percentage, tail DNA percentage, tail length, and tail moment in a concentration-dependent manner. The current results showed that SA, SAPP, and CA are cytotoxic and genotoxic to isolated lymphocytes in vitro.

Effects of the food additives sodium acid pyrophosphate, sodium acetate, and citric acid on hemato-immunological pathological biomarkers in rats: Relation to PPAR-α, PPAR-γ and tnfα signaling pathway.

The food additives sodium acid pyrophosphate (SAPP), sodium acetate (SA), and citric acid (CA) were evaluated for their hemato-immunotoxic effects. Forty adult Sprague-Dawley rats were distributed into four groups and were orally administered water, SAPP (12.6 mg/kg), CA (180 mg/kg), or SA (13.5 mg /kg) daily for 90 days. Erythrogram and leukogram profiles were evaluated. The levels of lysozyme, nitric oxide, immunoglobulin, and phagocytic activity were measured. Histologic and immunohistochemical evaluations of splenic tissues were performed. Changes in the mRNA expression levels of peroxisome proliferator-activated receptor α and γ (PPAR-α and PPAR-γ), and tumor necrosis factor α (TNF-α) genes were assessed. A significant leukopenic condition was observed with SAPP, while CA induced marked leukocytosis, and SA showed a lymphocytosis condition. Both the innate and humoral parameters were significantly depressed. Various pathological lesions were observed, including diffuse hyperplasia of the red pulp, depletion of the white pulp, and capsular and parenchymal fibrosis. A marked decrease in CD3 T-lymphocyte and CD20 B-lymphocyte immunolabeling in rats treated with SAPP and SA was evident. Marked downregulation of PPAR-α and PPAR-γ together with upregulation of TNF-α was recorded. These results indicate that high doses of SAPP, SA and CA exert hematotoxic and immunotoxic effects with long-term exposure.

Neighbourhood fluorosis in people residing in the vicinity of superphosphate fertilizer plants near Udaipur city of Rajasthan (India).

Chronic industrial fluoride toxicosis in the forms of dental, skeletal and non-skeletal fluorosis was investigated in 162 villagers (94 males and 78 females) above 15 years of age living in the vicinity of superphosphate fertilizer plants located approximately 12 km south of Udaipur city of Rajasthan, India. Out of these villagers, 90 (55.5%) and 29 (18.0%) were found to be afflicted with mild to severe dental and skeletal fluorosis, respectively. Dental fluorosis characterized with light to deep-brownish bilaterally striated horizontal lines, pits or patches and fine dots or granules was noted on incisor teeth of villagers. Irregular wearing, excessive corrosions (abrasions), dark-brownish pigmentation of exposed cementum and dentine material, diastem as between teeth, pronounced loss of tooth supporting bone with recession and bulging of gingiva (gum) were also present in subjects of older age group (>55 years). Among 29 (18.0%) individuals, mild to moderate manifestations of skeletal fluorosis such as crippling, kyphosis, invalidism and genu-varum syndrome were found. In these fluorotic subjects pain/rigidity in major joints viz. neck, back, hip, knee and shoulder was also found. None of the fluorotic subjects showed evidence of genu-valgum syndrome. Other signs of chronic industrial fluoride intoxication in soft tissues (non-skeletal fluorosis) included colic, intermittent diarrhoea or constipation, bloating, polyuria and polydipsia. These findings indicate that surrounding environment of superphosphate fertilizer plants is contaminated with fluoride emission, which in turn is causing diverse ill health effects in humans which are discussed.

Phosphocalyculin C as a pyrophosphate protoxin of calyculin C in the marine sponge Discodermia calyx.

Calyculin C, a minor derivative of the calyculins, has an additional methyl group on C32 of calyculin A. A recent biosynthetic study of calyculins revealed that an end product of calyculin biosynthesis is the pyrophosphate form, phosphocalyculin A. However, the pyrophosphate counterpart derived from calyculin C had not been reported. We isolated phosphocalyculin C as a minor pyrophosphate derivative, by a detailed investigation of an extract from the sponge Discodermia calyx. The treatment of phosphocalyculin C with the D. calyx cell-free extract significantly enhanced its cytotoxicity, providing molecular evidence for its role as the protoxin of calyculin C.

The cytotoxic effects of three different bisphosphonates in-vitro on human gingival fibroblasts, osteoblasts and osteogenic sarcoma cells.

INTRODUCTION: Osteonecrosis of the jaw (ONJ) is an emerging condition in patients undergoing long-term administration of bisphosphonates (BP) for the treatment of osteoporosis and hypercalcaemia associated with malignancy, multiple myeloma, and metastatic breast and prostate cancers. This is a follow-up study, its purpose was to examine the effects in-vitro of intravenous zoledronic acid (ZOL) and pamidronate (PAM) and oral alendronate (FOS) on the human oral cavity using gingival fibroblasts and osteoblasts cells and, in addition, osteogenic sarcoma cells (SaOS-2-cells). MATERIALS AND METHODS: Human gingival fibroblasts, osteoblasts and SaOS-2-cells were seeded on multiple 6-well plates at a density of 5 × 10(5)cells in a 4-week cell culture. Four different concentrations (1, 5, 10, 20 µM) of each BP (ZOL, PAM, FOS) and pyrophosphate were used in this study. RESULTS: All BP decreased collagen production and lowered cell proliferation in-vitro. ZOL was the component with most inhibitory effect. CONCLUSION: The findings in this study suggest that ZOL, PAM and FOS generally diminish cell proliferation and collagen production of human gingival fibroblasts, osteoblasts and SaOS-2-cells. The present follow-up study shows that not only ZOL and PAM but also FOS have a strong inhibitory effect on collagen production and cell survival in-vitro.

Inorganic materials as ameliorants for soil remediation of metal toxicity to wild mustard (Sinapis arvensis L.).

The ameliorating effects of different inorganic materials were investigated on a soil originating from a zinc smelter dumping site contaminated by toxic metals. Wild mustard (Sinapis arvensis L.) was used as a test plant. The soil was amended with different doses of mining sludge, Perferric Red Latosol (LVj), steel shots, cyclonic ash, silifertil, and superphosphate. The most effective amendments improved plant growth with 45% and reduced metal uptake by over 70% in comparison to untreated soil. Reductions in availability as estimated by BaCl2-extractable metals reached up to 90% for Zn and 65% for Cd as compared to unamended soil. These reductions were associated with lower shoot and root metal contents. Shoot Zn content was reduced from 1,369 microg g(-1) in plants grown on untreated soil to 377 microg g(-1) when grown on cyclonic ash amended soil while Cd decreased from 267 to 44 microg g(-1) in steel shots amended soil. Superphosphate addition had no ameliorating effect. On the contrary, it increased BaCl2-extractable amounts of Zn. Considering all parameters we determined, steel shots, cyclonic ash and silifertil are the most promising for remediating metal contaminated soil in the tropics. Further studies evaluating impacts, cost-effectiveness and durability of effects will be conducted.

Mutagenicity of octane and tetrasodium pyrophosphate in bacterial reverse mutation (Ames) test.

We investigated the genotoxicities or mutagenicities of 2 chemicals (octane and tetrasodium pyrophosphate) with limited toxicological data in spite of their common usage based on Ames reverse mutation test. In this test, treatment of 2 chemicals at each five dose did not induce mutagenicity in Salmonella typhimurium TA98, TA100, TA1535, TA1537, and in Escherichia coli WP2uvrA with and without metabolic activation. These results indicate that 2 chemicals do not have mutagenic potentials under the conditions examined in each study. Despite these results, it can affect by inducing inhalation, skin or eye contact, ingestion, and have affected central nervous system as a target organ. It is thus necessary to prepare the local exhaust system and personal protective equipments. Based on this study, we suggest that future studies should be directed toward chronic inhalation, carcinogenic test and so on.

Bioassay and use in irrigation of untreated and treated wastewaters from phosphate fertilizer industry.

Wastewater from phosphate fertilizer industry that contains essentially a significant amount of both fluoride and phosphate was treated by separative precipitation of fluoride ions with hydrated lime. Thus, a phosphate-rich effluent with low content of fluoride was obtained. The microtoxicity of the treated wastewater was then monitored by LUMIStox and its phytotoxicity was investigated on tomato (Lycopersicon esculentum), wheat (Triticum aestivum), maize (Zea mays), ryegrass (Lolium perenne), and alfalfa (Medicago sativa) seed germination and plant growth. The cress (Lepidium sativum) was used as a standard species for the germination index and phytotoxicity evaluation. Seedlings of four species (namely wheat, maize, ryegrass, and alfalfa) were grown in pots, which were irrigated with untreated wastewater, treated wastewater, aqueous solution of triple superphosphate fertilizer (TSP) or with tap water as control. LUMIStox tests showed that lime treatment allowed a significant toxicity removal. The treated water displayed beneficial fertilizing effect on plants. An increase in the germination index from 100% to 119% was observed. However, the untreated wastewater inhibited the species germination even when diluted 10 times. Neither plants mortality nor growth inhibition was observed after 90 days of treated wastewater application. Moreover, an improvement in plant growth, leaf number and a root development were noticed in these plants when compared with those irrigated with tap water or with fertilizer. In contrast, leaf necrosis and growth inhibition were observed in plants amended with raw wastewater. The irrigation with treated wastewater also improved soil labile P content. Indeed, soils amended with treated wastewater had more a double labile P concentration (38.15 mg kg(-1)) in comparison with control soil (15.53 mg kg(-1)).

Vascular targeting agent Oxi4503 inhibits tumor growth in a colorectal liver metastases model.

BACKGROUND AND AIM: Oxi4503 is a potent vascular targeting agent belonging to the family of combretastatins. These agents produce an acute reduction in tumor blood flow leading to tumor necrosis. Despite evidence of its efficacy in certain malignancies, the effect on colorectal liver metastases remains largely unknown. This study investigates the effect of Oxi4503 on colorectal liver metastases in a murine model. METHODS: The effect of a single dose of Oxi4503 on established tumors in a murine model of colorectal liver metastases was assessed following administration of 1-50 mg/kg Oxi4503. In addition, the effects of continuous, daily and intermittent dosing (1-5 mg/kg) on tumor necrosis and growth were studied by quantitative histological and stereological analysis. The effect of multiple dosing on long-term survival was also assessed using the Kaplan-Meier analysis. The microvascular effects of therapy were studied by scanning electron microscopy of microvascular resin casts. RESULTS: A single dose of 5 or 50 mg/kg of Oxi4503 produced significant tumor necrosis compared to the controls. Subcutaneous continuous dosing infusion with Oxi4503 at 1 mg/kg/day reduced tumor growth compared to the controls, but was associated with marked systemic toxicity. Daily administration over 21 days was associated with significant mortality. Intermittent dosing of Oxi4503 (two doses, 3 days apart) produced the greatest reduction in tumor growth with minimal toxicity and conferred a significant survival advantage. Microvascular casts demonstrated significant disruption of tumor vessels. CONCLUSIONS: A single dose of Oxi4503 produced significant necrosis and microvascular injury in colorectal liver metastases. Intermittent dosing with Oxi4503 produced the maximum reduction in tumor growth, minimal toxicity, and a significant improvement in survival. Oxi4503 is a potential anticancer agent. Further research into its mechanism of action and its synergistic use with other therapies is warranted.

Effect of vascular targeting agent Oxi4503 on tumor cell kinetics in a mouse model of colorectal liver metastasis.

BACKGROUND: Oxi4503 has been shown to inhibit tumor growth and improve survival in an animal model of colorectal (CRC) liver metastases. This agent appears to selectively target the endothelial cytoskeleton with resultant vessel occlusion and tumor necrosis. MATERIALS AND METHODS: This study evaluated the pattern of tumor necrosis caused by Oxi4503, with particular emphasis on patterns of cell proliferation and apoptosis in a murine model of CRC liver metastases. RESULTS: A single dose of Oxi4503 caused immediate tumor vasculature collapse and subsequently tumor necrosis. There was widespread central necrosis with evidence of viable tumor cells at the periphery. Alterations in the number and spatial pattern of tumor cells undergoing apoptosis and the rate of cellular proliferation were also observed following treatment. Microvessel density was reduced following treatment, however patent vessels were still observed within the necrotic core. CONCLUSION: Although Oxi4503 caused significant tumor destruction, synergistic treatment with cytotoxic and/or anti-angiogenic agents should be considered in order to achieve complete tumor eradication and long-term survival.

Effects of malathion plus superphosphate or urea on Najdi sheep.

The effects of p.o. administration of the combination of malathion + superphosphate or urea on Najdi sheep were evaluated in sheep dosed as untreated controls, malathion-treated at 25 mg/kg/d, superphosphate-treated at 450 mg/kg/d, urea-treated at 450 mg/kg/d, malathion-treated at 25 mg/kg/d + superphosphate treated at 450 mg/kg/d, or malathion treated at 25 mg/kg/d + urea treated at 450 mg/kg/d. Oral doses of malathion alone were lethal after 6 d, and malathion + urea were fatal after 6-8 d. Malathion + superphosphate caused death after 2-3 d. Malathion, but not superphosphate or urea, inhibited serum cholinesterase activity. Hepatonephropathy correlated with changes in serum AST, ALP, cholesterol, triglyceride, bilirubin, urea, total protein and albumin. Neither malathion nor its combination with superphosphate or urea caused peripheral neuropathy.

Cytotoxicity of modern dentin adhesives--in vitro testing on gingival fibroblasts.

The present investigation was designed to test cellular toxicity of modern dentin adhesives. With the use of the products Ariston Liner, Etch & Prime 3.0, Optibond Solo, Prime & Bond NT, Scotchbond 1, and Syntac Sprint, test specimens were prepared according to the manufacturers' instructions and transferred into a culture medium. Eluates were obtained and pipetted onto fibroblast cultures, incubated, and subsequently stained. The respective cell densities and the numbers of normal, altered, and dead cells were determined and compared with control cell cultures. Statistical analysis of the data showed that all materials caused cytotoxic effects. Scotchbond 1 displayed the highest number of dead cells. The difference was statistically significant compared to Etch" 3.0, Optibond Solo, Prime&Bond NT, and the control. The lowest cell density was found for Scotchbond 1 and Ariston Liner. The difference was also statistically significant in comparison with Etch" 3.0, Optibond Solo, Prime&Bond NT, and the control. To conclude, all tested dentin adhesives caused cytotoxic reactions. Taking the limitations of an in vitro experiment into consideration, Prime&Bond NT, Optibond Solo, and Etch" 3.0 appear to be the most recommendable products, and Scotchbond 1 and Ariston Liner the least.

[The hygienic assessment of superphosphates manufactured on a base of Algerian phosphorites]. / Gigienicheskaia otsenka superfosfatov, izgotovlennykh na osnove alzhirskikh fosforitov.

Hygienic and toxicological investigations of soil, plants, and animals have shown that the superphosphates made from Algerian phosphorites little differ from those made from the apatites of the Kola Peninsula. Superphosphates A and B should be referred to as hazard class IV and the superphosphates treated by ammonium should be classified as hazard class III.

Mechanical properties and histological evaluation of sintered beta-Ca2P2O7 with Na4P2O7.10H2O addition.

The ultimate goal of implantation of biomaterials in the skeleton is to reach full integration of the non-living implant with the living bone. The biomaterial can be used much as a bone graft, resorbing or dissolving as bone growth occurs, and the end result is a new remoulded bone. Calcium pyrophosphate, Ca2P2O7, is one of the intermediate products of bone mineralization. beta-Dicalcium pyrophosphate (beta-DCP) doped with certain amounts of Na4P2O7.10H2O was prepared as the developed material. Na4P2O7.10H2O was used as a liquid-phase additive to improve the sintering process and promote physiological bioresorbability. Compressive strength and four-point bending strength were measured by the Bionix test system 858. The mechanical strength of the sintered beta-DCP increased with the addition of Na4P2O7.10H2O up to 5 wt%, but thereafter decreased. The microstructure and crystal structure were analysed by the techniques of SEM, EPMA, TEM and XRD. The relationship between the mechanical strength of the sintered bioceramics and the Na4P2O7.10H2O dopant was examined in terms of the presence of NaCa(PO3)3, grain growth and abnormal grain coalescence while the dopant increased. Preliminary in vivo evaluation was studied by rabbit femur condyle implantation. There was no inflammation or any toxic sign during the experimental period. The histological section of intraosseous implantation revealed that the new bone deposited directly on the surface of the material in the fourth week after operation. The implant gradually decreased in volume and was replaced by the surrounding regenerated bone in the rabbit condyle in vivo environment. The results led us to conclude that the developed material has great potential as a biodegradable bone substitute.

[Study of the mutagenic properties of oligonucleotides containing and internucleotide pyrophosphate bond using a mutation system based on phage M13]. / Issledovanie mutagennykh svoistv oligonukleotidov, soderzhashchikh mezhnukleotidnoe pirofosfatnoe zveno, c pomoshch'iu mutatsionnoi sistemy na osnove faga M13.

Mutagenic properties of oligonucleotides with pyrophosphate internucleotide bond was studied. It was shown that the pyrophosphate bond in the oligo structure does not induce mutations but promotes a more efficient induction of marker deletions predetermined by the nucleotide sequence as compared to the native oligonucleotide. Marker deletion induction proceeds according to the repair mechanism as homozygotes dominate in the mutant generation.

Randomness of chromosome breaks in bone marrow cells of fertilizer-fed mice, Mus musculus.

Three commonly used fertilizers, urea, single superphosphate and muriate of potash, induced chromosome and chromatid breaks in the metaphase chromosomes of bone marrow cells of fertilizer-fed Swiss albino mice, Mus musculus. The breaks caused by urea and phosphate were non-randomly distributed, since they were more frequent in the longer chromosomes than in the smaller ones, and more common in the distal region than in the juxtacentromeric and median regions. The breaks induced by muriate of potash were randomly distributed in both the length and region of the chromosomes.

[The effect of mineral fertilizers on mosquito larvae (Culicidae)]. / Vliianie mineral'nykh udobrenii na lichinok komarov (Culicidae).

The effect of superphosphate, nitrophoska and ammophos on larvae of Culicidae was studied under laboratory and field conditions of Novgorod Province. Superphosphate better than other fertilizers acidifies water, while nitrophoska and ammophos increase the ammonia contents of 27 to 240 fold that affects lethally the larvae. 0.4-0.5% and 1% concentrations of mineral fertilizers are a lethal dose for I-II instar and III-IV instar larvae, respectively.