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PubMed2002

Effect of preservatives on IgG aggregation, complement-activating effect and hypotensive activity of horse polyvalent antivenom used in snakebite envenomation.

Intravenous administration of antivenoms is associated with early adverse reactions in a number of cases, but the causes of this phenomenon are still unclear. The effect of preservatives (phenol and thimerosal) on IgG aggregate and dimer formation, in vitro complement-activating effect and hypotensive activity of a whole IgG horse liquid polyvalent antivenom, produced by caprylic acid fractionation, was assessed. These parameters were studied since they have been associated with the development of early adverse reactions to the administration of antivenoms and human immunoglobulins. After a three-year storage period at 4 degrees C, antivenoms with preservatives had an increased content of IgG aggregates and dimers when compared with antivenom devoid of phenol and thimerosal. These observations correlate with a slight increment in the turbidity of preservative-containing antivenoms. The three antivenoms studied (formulation: no preservatives; with phenol and thimerosal; with thimerosal alone) activated human complement in vitro, with only minor quantitative differences among them. When antivenoms were administered as a bolus intravenous injection in rats, a rapid and prominent hypotension of short duration was observed after injection of phenol-containing antivenom, whereas such an effect was absent in antivenom free of preservative and in the one containing only thimerosal. Bolus injection of saline solution with phenol resulted in a similar hypotension, indicating that the effect is due to phenol. However, when phenol-containing antivenom was diluted 1:5 with saline solution before infusion, as occurs in the clinical use of this product, no hypotension was observed. Our results stress the need to evaluate the effects of preservatives on the physicochemical and pharmacological characteristics of antivenoms.

Animals↗
PubMed1993

C-mitosis and numerical chromosome aberration analyses in human lymphocytes: 10 known or suspected spindle poisons.

As a part of a coordinated EEC project to validate suitable assays for chemically induced genomic mutations, numerical chromosomal aberrations and spindle effects were studied in human lymphocyte cultures exposed to cadmium chloride, chloral hydrate, colchicine, diazepam, econazole, hydroquinone, pyrimethamine, thiabendazole, thimerosal and vinblastine. Chromosome number analysis was carried out after treatment for 48 and 72 h; spindle effects, i.e., increases in the mitotic indices and c-mitoses, were analyzed in cultures treated 5 h before fixation. Dose-related numerical chromosomal aberrations are induced by colchicine and vinblastine, the only chemicals that also induce c-mitotic effects in a wide range of doses. Hyperdiploidy is induced by chloral hydrate, cadmium chloride and thimerosal without dose-effect relationship; chloral hydrate and thimerosal affect spindle functions while only a weak spindle effect is produced by cadmium chloride. Tetraploid and/or endoreduplicated cells are induced without dose-effect relationship by hydroquinone, thiabendazole and thimerosal, all of them able to produce c-mitotic effects. Diazepam and econazole induce only hypodiploidy; pyrimethamine does not induce numerical chromosomal aberrations.

Cadmium↗
PubMedDate not supplied

Allergic and toxic reactions of soft contact lens wearers.

The allergic and toxic effects of preservatives found in soft contact lens solutions are reviewed. Thimerosal, a preservative commonly found in soft contact lens solutions, may cause ocular delayed hypersensitivity. Patients with delayed hypersensitivity to thimerosal may develop conjunctival hyperemia, corneal infiltrates and intolerance to lens wear with the use of soft contact lens solutions containing thimerosal. Delayed hypersensitivity to thimerosal can be demonstrated by an occlusive patch test or intradermal injection. Discontinuation of chemical disinfection with substitution of thermal disinfection using unit-dose, non-preserved saline causes resolution of signs and symptoms. Another cause of lens wear intolerance is contact lens-associated giant papillary conjunctivitis. Protein deposits on the lens may act as antigens and initiate an allergic condition which disappears when the lens is discontinued. Alternatively, the giant papillae may be the result of mechanical irritation induced by the contact lenses. The chemical preservatives that are found in soft contact lens solutions cause epithelial toxicity when applied to isolated rabbit corneas. However, corneal toxicity toxicity in a clinical setting has not been demonstrated.

Animals↗
PubMed2005

Reversible activation of secretory phospholipase A2 by sulfhydryl reagents.

Secretory phospholipase A(2)s (sPLA(2)s) have been implicated in physiological and pathological events, but the regulatory mechanism(s) of their activities in cells remains to be solved. Previously, we reported that phenylarsine oxide (PAO), a sulfhydryl reagent, stimulated arachidonic acid (AA) release in rat pheochromocytoma PC12 cells. In this study, we examined the effects of thimerosal, another sulfhydryl reagent, to clarify the sulfhydryl modification and activation of sPLA(2) molecules in cells. Like PAO, thimerosal-stimulated AA release in an irreversible manner and the responses were not additive. Dithiol compounds such as dithiothreitol inhibited AA release from both the thimerosal- and the PAO-treated cells, and monothiol compounds (l-Cys and glutathione) decreased the thimerosal response. Both sulfhydryl reagents stimulated AA release from the HEK293T cells expressing human sPLA(2)X, and stimulated the sPLA(2) activities of bee venom sPLA(2) and the soluble fraction of sPLA(2)X-expressing cells. Our results suggest that the sPLA(2)s in cells are inactive and modification of disulfide bonds in the molecules can be a trigger of sPLA(2) activation in cells. Sulfhydryl reagents are useful tools for studying the regulatory mechanism(s) of sPLA(2) activity in cells.

Animals↗
PubMed1997

Mechanisms involved in ATP-evoked Ca2+ oscillations in isolated human granulosa-luteal cells.

Using single-cell microfluorimetry, we have shown that ATP evoked repetitive Ca2+ oscillations in intact Fura-2 loaded human granulosa-luteal cells (hG/LCs) in the absence of extracellular Ca2+. Sustained increases in [Ca2+]i required extracellular Ca2+ and ATP depleted stores were refreshed by brief (2 min) incubation with external Ca2+. Basal [Ca2+]i was unaffected by caffeine (1 mM), but 20 mM caffeine inhibited ATP-evoked Ca2+ release in the absence of external Ca2+. Thimerosal (10 microM) evoked repetitive Ca2+ spikes, under Ca(2+)-free conditions, which fused to form an elevated plateau when external Ca2+ was replaced. Thimerosal-induced changes in [Ca2+]i were reversibly inhibited by the thiol reducing agent dithiothreitol (1 mM). The periodicity and amplitude of the [Ca2+]i oscillations produced by thimerosal and ATP differ. ATP- or thimerosal-evoked changes in [Ca2+]i were unaffected by dantrolene sodium (10 microM). The Ca(2+)-ATPase inhibitor thapsigargin (1 microM) increased [Ca2+]i and attenuated subsequent ATP-evoked changes in [Ca2+]i. We conclude that ATP stimulates an oscillatory release of Ca2+ from InsP3-sensitive stores in hG/LCs.

Adenosine Triphosphate↗
PubMed2000

Regulation of sodium currents through oxidation and reduction of thiol residues.

Changes in redox state are involved in several physiological and pathophysiological processes. Previous experiments have demonstrated that nitric oxide can function as a reactive oxygen species, inhibiting neuronal sodium currents by nitrosylation of thiol residues. We hypothesized that nitric oxide and thiol oxidizers similarly modulate voltage-dependent sodium currents. Voltage-dependent sodium currents were studied with the whole-cell patch-clamp technique in NB41A3 neuroblastoma cells. The nitric oxide donor 3-(2-hydroxy-2-nitroso-1-propylhydrazino)-1-propanamine did not affect sodium currents. In contrast, the thiol oxidizers thimerosal and 4,4'-dithiopyridine significantly inhibited sodium currents. The effect of thimerosal persisted after washout, but could be fully reversed by the reducing agent dithiothreitol. Reduced glutathione did not restore the sodium current amplitude when given extracellularly, while intracellular glutathione prevented the inhibitory effect of thimerosal. Pretreatment with the alkylating agent N-ethylmaleimide blocked the inhibitory action of thimerosal. Thiol oxidation caused a shift in the voltage dependence of fast and slow inactivation to more hyperpolarized potentials without concomitant effects on the voltage dependence of activation. Mercaptoethanol and reduced glutathione enhanced sodium currents by shifting the voltage dependence of inactivation to depolarized potentials. These results demonstrate that the oxidation and reduction of thiol residues alters the properties of voltage-sensitive sodium channels and may play an important role in the regulation of membrane excitability.

Animals↗
PubMed2002

Irregular spiking in free calcium concentration in single, human platelets. Regulation by modulation of the inositol trisphosphate receptors.

Fluorescence ratio imaging indicates that immobilized, aspirin-treated platelets, loaded with Fura-2, respond to inositol 1,4,5-trisphosphate- (InsP3)-generating agonists such as thrombin by high-frequency, irregular rises in cytosolic [Ca2+]i with spikes that vary in peak level and peak-to-peak interval. This differs from the regular [Ca2+]i oscillations observed in other, larger cells. We found that the thiol-reactive compounds thimerosal (10 microm) and U73122 (10 microm) evoked similar irregular Ca2+ responses in platelets, but in this case in the absence of InsP3 generation. Thrombin-induced spiking was acutely abolished by inhibiting phospholipase C or elevating intracellular cAMP levels, while spiking with sulfhydryl reagents was only partially blocked by cAMP elevation. Confocal laser scanning microscopy using fluo-3-loaded platelets indicated that, with all agonists or conditions, the irregular spikes were almost instantaneously raised in various regions within a single platelet. When using saponin-permeabilized platelets, we found that InsP3-induced Ca2+ release from stores was stimulated by modest Ca2+ concentrations, pointing to a mechanism of InsP3-dependent Ca2+-induced Ca2+ release (CICR). This process was completely inhibitable by heparin. The Ca2+ release by InsP3, but not the CICR sensor, was negatively regulated by cAMP elevation. Thimerosal treatment did not release Ca2+ from intracellular stores, but markedly potentiated the stimulatory effect of InsP3. In contrast, U73122 caused a heparin/cAMP-insensitive Ca2+ leak from stores that differed from those used by InsP3. Taken together, these results demonstrate that InsP3 receptor channels play a crucial role in the irregular, spiking Ca2+ signal of intact platelets, even when induced by agents such as thimerosal or U73122 which do not stimulate InsP3 formation. The irregular Ca2+ release events appear to be subjected to extensive regulation by: (a) InsP3 level, (b) the potentiating effect of elevated Ca2+ on InsP3 action via CICR, (c) InsP3 channel sensitization by sulfhydryl (thimerosal) modification, (d) InsP3 channel-independent Ca2+ leak with U73122, and (e) down-regulation via cAMP elevation. The observation that individual Ca2+ peaks were generated in various parts of a platelet at similar intervals and amplitudes points to effective cooperation of the various stores in the Ca2+-release process.

Alprostadil↗
PubMed1995

The effect of mersalyl on inositol trisphosphate receptor binding and ion channel function.

A number of thiol-reactive agents induce repetitive Ca2+ spiking in cells by a mechanism thought to involve sensitization of the inositol 1,4,5-trisphosphate receptor (IP3R). To further define the basis of this interaction, we have studied the effect of several thiol-reactive agents on [3H]IP3 binding, IP3-gated channel activity, and conformation of the IP3R in membranes from hepatocytes, cultured WB rat liver epithelial cells, and cerebellum microsomes. At 4 degrees C, the organomercurial thiol-reactive agent mersalyl markedly stimulates (3-4fold) [3H]IP3 binding to permeabilized hepatocytes. The closely related molecule, thimerosal, has only a small stimulatory effect under these conditions, and GSSG or N-ethylmaleimide are without effect. The stimulatory effect of mersalyl was associated with a decrease in Kd of the IP3R with no change in Bmax. Mersalyl was without effect on detergent-solubilized hepatocyte binding sites or on the [3H]IP3 binding activity of cerebellum microsomes. In contrast to thimerosal, which potentiates IP3-mediated Ca2+ release, mersalyl blocked IP3-gated Ca2+ channels. Mersalyl pretreatment of WB membranes altered the pattern of immunoreactive receptor fragments generated upon subsequent cleavage of the receptor with proteinase K. This effect was not reproduced by thimerosal and was also not observed in experiments on cerebellum microsomes. We conclude that the WB cell and brain IP3 receptors are differently regulated by modification of thiol groups. Reaction of the WB cell IP3 receptor with mersalyl alters its conformation and modifies the accessibility of sites on the protein that are cleaved by proteinase K. In the presence of mersalyl, the receptor has high affinity for IP3 but is inactive as a Ca2+ channel. This contrasts with the high affinity receptor/active Ca2+ channel induced by thimerosal, suggesting that even closely related thiol agents may interact at different thiol groups.

Animals↗
PubMed1994

Regulation of intracellular calcium in the mouse egg: calcium release in response to sperm or inositol trisphosphate is enhanced after meiotic maturation.

Fertilization of the immature, prophase I-arrested mouse oocyte produces multiple Ca2+ transients similar to those of the mature, metaphase II egg; however, the first Ca2+ transient is much lower in amplitude and shorter in duration. In contrast to prophase I-arrested oocytes, maturing oocytes fertilized after germinal vesicle breakdown have first Ca2+ transients similar to those of mature fertilized eggs. Immature, prophase-arrested oocytes release less Ca2+ in response to injection of inositol 1,4,5-trisphosphate (IP3) than eggs. At high concentrations, the sulfhydryl reagent, thimerosal (200 microM), causes Ca2+ oscillations in eggs and produces similar oscillations in oocytes. A lower concentration of thimerosal (25 microM) does not cause Ca2+ oscillations, but does sensitize IP3-induced Ca2+ release in both eggs and oocytes, since IP3-induced Ca2+ release is enhanced in the presence of 25 microM thimerosal. Incubation of oocytes in 25 microM thimerosal before injection of 2.2 microM IP3 causes oocytes to release as much Ca2+ as is released in eggs injected with 2.2 microM IP3. These results indicate that immature mouse oocytes possess intracellular stores of releasable Ca2+ similar in size to Ca2+ stores in eggs; however, these stores are less sensitive to IP3. Development of the IP3-induced Ca2+ release mechanism may be an important component of maturation; at fertilization of the egg, Ca2+ must be elevated to levels sufficient to activate further development and establish a block to polyspermy. Mouse oocytes appear to develop an increased sensitivity to IP3 during the course of oocyte maturation.

Animals↗
PubMed2000

Mechanism of intracellular pH increase during parthenogenetic activation of In vitro matured porcine oocytes.

Parthenogenetic activation of porcine oocytes by using 7% ethanol, 50 or 100 microM A23187 results in an increase in intracellular pH as does prolonged exposure to thimerosal. We attempt to specify which transporters or mechanisms are involved in the observed increase in intracellular pH during oocyte activation. Experiments were performed in the absence of sodium; the presence of 2.5 mM amiloride, a potent inhibitor of the Na(+)/H(+) antiport; in the absence of bicarbonate; and in the presence of 4, 4'-diisothiocyanatodihydrostilbene-2,2'-di-sulfonic acid, disodium salt (H(2)DIDS) for all three activation methods. These treatments had no effect on the increase in intracellular pH induced by the calcium ionophore or thimerosal, but all reduced the increase in pH (P < 0.001) in the 7% ethanol group. This suggests that the Na(+)/H(+) antiport and the HCO(3)(-)/Cl(-) exchangers are not playing a role during treatment with calcium ionophore or thimerosal, and the pH increase observed during treatment with 7% ethanol may be dependent upon a sodium or bicarbonate flux (or both) into the oocyte. Bafilomycin A1 (500 nm), an inhibitor of vacuolar-type H(+) ATPases, had no effect on 7% ethanol or thimerosal treatments, but significantly reduced the increase in intracellular pH observed during calcium ionophore treatment. This may be the result of an initial local increase in intracellular free calcium levels.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed2005

Contact sensitivity to preservatives in Turkey.

This study was designed to evaluate the frequency of contact sensitivity to 14 common preservatives among patients with contact dermatitis in Turkey. From 2000 to 2004, 308 patients with the diagnosis of contact dermatitis were patch tested in the Department of Dermatology, Ankara University School of Medicine. All patients were patch tested with European standard series. In addition to the four preservatives included in the standard series, patients were also tested with DMDM hydantoin, imidazolidinyl urea, bromonitropropane diol, diazolidinyl urea, thimerosal, propylene glycol, chlorocresol, chloroxylenol, methyldibromoglutaronitrile/phenoxyethanol (MDBGN/PE) and benzalkonium chloride. Out of the 308 patients suspected of having contact dermatitis, 23 patients were found to have positive reactions to one or more preservatives. Preservatives that were the most frequent cause of positive reactions were thimerosal (1.6%), benzalkonium chloride (1.6%), formaldehyde (1.3%) and MDBGN/PE (0.9%). In our study, 65% of the positive reactions were caused by allergens not present in the standard series, such as thimerosal, benzalkonium chloride and MDBGN/PE. Although thimerosal caused a high rate of contact sensitivity, it may not be considered as an important allergen, because clinical relevance could not be found in any of the patients.

Adolescent↗
PubMed1989

Cytotoxicity and mutagenicity of ophthalmic solution preservatives and UVA radiation in L5178Y cells.

Four chemical preservatives commonly used in ophthalmic solutions were tested for their toxic and mutagenic potential in mouse lymphoma cells with and without exposure of the cells to ultraviolet A (UVA) radiation. The preservatives tested were benzalkonium chloride (BAK), chlorhexidine, thimerosal and ethylenediaminetetraacetic acid (EDTA). Cell survival and mutagenesis were measured using the L5178Y mouse lymphoma (TK +/-) system. Cells were exposed to varying amounts of preservatives for 1 h at 37 degrees C, and then aliquots were irradiated with UVA radiation (during the exposure to preservative). Cells were then assayed for survival, and for mutagenesis at the thymidine kinase (TK) locus. In concentrations commonly found in ophthalmic solutions, BAK, chlorhexidine, and thimerosal were toxic to cells, and thimerosal was slightly mutagenic. When cells were exposed to preservative and UVA radiation, chlorhexidine was mutagenic and the mutagenic activity of thimerosal was enhanced.

Animals↗
PubMed1995

Activation of store-operated calcium influx at resting InsP3 levels by sensitization of the InsP3 receptor in rat basophilic leukaemia cells.

1. Patch clamp and fura-2 AM measurements were performed to study the effects of sensitizing the inositol 1,4,5-trisphosphate (InsP3) receptor to InsP3 on the activation of Ca2+ release-activated Ca2+ current (ICRAC) in rat basophilic leukaemia (RBL) cells. 2. The sensitizing agent thimerosal (1 microM) triggered Ca2+ release, and this was followed by Ca2+ influx. With no added InsP3 in the patch pipette, thimerosal activated ICRAC; this was prevented by heparin. ICRAC activated by thimerosal was very similar to that evoked by InsP3 or ionomycin. 3. Dialysing cells either for short (30 s) or long (600 s) periods of time prior to application of thimerosal did not affect the subsequent activation of ICRAC, even though no InsP3 was included in the patch pipette. 4. These results suggest that sensitizing the InsP3 receptor can result in large Ca2+ influx in the presence of resting InsP3, and that stores closer to the membrane may contribute more to activation of ICRAC than stores further away.

Animals↗
PubMed2003

Mechanism of inhibitory actions of oxidizing agents on calcium-activated potassium current in cultured pigment epithelial cells of the human retina.

PURPOSE: To identify the mechanisms by which oxidative stress with oxidizing agents alters the activity of ion channels in human retinal pigment epithelial (RPE) cells. METHODS: The effects of oxidizing agents on ion currents were investigated in human RPE R-50 cells with the aid of the whole-cell, cell-attached, and inside-out configurations of the patch-clamp technique. RESULTS: In the whole-cell configuration, t-butyl hydroperoxide (t-BHP; 1 mM), thimerosal (30 microM), and 4,4'-dithiodipyridine (DTDP; 30 microM) suppressed voltage-dependent K(+) current (I(K)) that was sensitive to inhibition by iberiotoxin or paxillin, yet not by apamin or 5-hydroxydecanoate sodium. Meclofenamic acid or Evans blue, but not diazoxide, reversed the decrease in I(K) caused by t-BHP. In cells dialyzed with ceramide (30 microM), neither t-BHP (1 mM) nor thimerosal (30 microM) had any effect on I(K), whereas DTDP (30 microM) slightly suppressed it. In cell-attached recordings, t-BHP (1 mM), thimerosal (30 microM), and DTDP (30 microM) suppressed the activity of large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels. Dithiothreitol (10 microM) reversed DTDP-induced decrease in channel activity. Under current-clamp conditions, cell exposure to oxidizing reagents caused membrane depolarization. In cells dialyzed with ceramide (30 microM), membrane potential remained unaltered in the presence of t-BHP. CONCLUSIONS: The results demonstrate that hydrophilic oxidants (e.g., t-BHP and thimerosal) suppress I(K) and suggest that the underlying mechanism of this inhibitory action may involve the generation of intracellular ceramide. However, the inhibition of BK(Ca) channels by DTDP, a membrane-permeable oxidant, in human RPE cells may result from the direct inhibition of BK(Ca) channels and indirectly from an increase in the intracellular production of ceramide.

Cells, Cultured↗
PubMed2006

A meta-analysis epidemiological assessment of neurodevelopmental disorders following vaccines administered from 1994 through 2000 in the United States.

BACKGROUND: Thimerosal is an ethylmercury-containing compound (49.6% mercury by weight) used as at the preservative level in vaccines (0.005% to 0.01%). METHODS: Statistical modeling in a meta-analysis epidemiological assessment of the Vaccine Adverse Event Reporting System (VAERS) for neurodevelopment disorders (NDs) reported following Diphtheria-Tetanus-whole-cell-Pertussis (DTP) vaccines in comparison to Diphtheria-Tetanus-whole-cell-Pertussis-Haemophilus Influenzae Type b (DTPH) vaccines (administered: 1994-1997) and following Thimerosal-containing Diphtheria-Tetanus-acellular-Pertussis (DTaP), vaccines in comparison to Thimerosal-free DTaP vaccines (administered: 1997-2000), was undertaken. RESULTS: Significantly increased adjusted (sex, age, vaccine type, vaccine manufacturer) risks of autism, speech disorders, mental retardation, personality disorders, thinking abnormalities, ataxia, and NDs in general, with minimal systematic error or confounding, were associated with TCV exposure. CONCLUSION: It is clear from the results of the present epidemiological study and other recently published data associating mercury exposure with childhood NDs, additional ND research should be undertaken in the context of evaluating mercury-associated exposures, especially from Thimerosal-containing vaccines.

Child, Preschool↗
PubMed1982

Ocular inflammation in patients using soft contact lenses.

Soft contact lenses cleaned and stored in thimerosal-containing solutions may produce an ocular inflammatory process and, if untreated, corneal neovascularization. We examined 38 patients who were using soft contact lenses and found that 31 patients had positive ocular provocation test results to a thimerosal-preserved artificial tears solution. Delayed hypersensitivity to thimerosal was demonstrated by positive patch tests in 27 patients. Avoidance of thimerosal and the use of unit-dose sterile saline solutions has resulted in complete resolution of ocular symptoms.

Adult↗
PubMed1999

Release of Ca(2+) from intracellular stores and entry of extracellular Ca(2+) are involved in sea squirt sperm activation.

A rise in intracellular free Ca(2+) concentration ([Ca(2+)](i)) is required to activate sperm of all organisms studied. Such elevation of [Ca(2+)](i) can occur either by influx of extracellular Ca(2+) or by release of Ca(2+) from intracellular stores. We have examined these sources of Ca(2+) in sperm from the sea squirt Ascidia ceratodes using mitochondrial translocation to evaluate activation and the Ca(2+)-sensitive dye fura-2 to monitor [Ca(2+)](i) by bulk spectrofluorometry. Sperm activation artificially evoked by incubation in high-pH seawater was inhibited by reducing seawater [Ca(2+)], as well as by the presence of high [K(+)](o) or the Ca channel blockers pimozide, penfluridol, or Ni(2+), but not nifedipine or Co(2+). The accompanying rise in [Ca(2+)](i) was also blocked by pimozide or penfluridol. These results indicate that activation produced by alkaline incubation involves opening of plasmalemmal voltage-dependent Ca channels and Ca(2+) entry to initiate mitochondrial translocation. Incubation in thimerosal or thapsigargin, but not ryanodine (even if combined with caffeine pretreatment), evoked sperm activation. Activation by thimerosal was insensitive to reduced external calcium and to Ca channel blockers. Sperm [Ca(2+)](i) increased upon incubation in high-pH or thimerosal-containing seawater, but only the high-pH-dependent elevation in [Ca(2+)](i) could be inhibited by pimozide or penfluridol. Treatment with the protonophore CCCP indicated that only a small percentage of sperm could release enough Ca(2+) from mitochondria to cause activation. Inositol 1,4,5-trisphosphate (IP(3)) delivered by liposomes or by permeabilization increased sperm activation. Both of these effects were blocked by heparin. We conclude that high external pH induces intracellular alkalization that directly or indirectly activates plasma membrane voltage-dependent Ca channels allowing entry of external Ca(2+) and that thimerosal stimulates release of Ca(2+) from IP(3)-sensitive intracellular stores.

Animals↗
PubMed1988

Broad reacting surface antigens in Leptospira biflexa serovar andamana.

Previous investigations had demonstrated that genus-specific and species-specific antigens of leptospires are deep-seated within the leptospiral cell. Conversely, the present study has shown that strain CH11, serovar andamana of the non-pathogenic species of Leptospira biflexa is endowed on its surface with two cross-reacting antigens; a newly recognized antigen common to L. interrogans and L. biflexa spp. and an antigenic determinant common to a previously described genus-specific protein antigen (GP-Ag). The serovar andamana, when thimerosal-treated, behaved like an interspecies-specific antigen, cross-reacting in the complement fixation test with sera from rabbits immunized with L. interrogans and L. biflexa spp. and with sera from subjects with leptospirosis from various serovars. In the immuno electron microscopic test, human leptospirotic sera bound to the surface of thimerosal-treated andamana and not to the surface of untreated andamana showing that an interspecies-specific antigen was located underneath the outermost layer of this serovar. In the same test, the monoclonal antibody GP-7 against the GP-Ag bound to the surface of untreated andamana and not to the surface of thimerosal-treated andamana, showing that an antigenic determinant, common to GP-Ag and different from the first one, was located on the outer membrane of andamana. In human leptospirotic sera, antibodies against the cross-reacting antigen of thimerosal-treated andamana demonstrated by the complement fixation test were formed earlier and in a higher percentage of sera than the serovar-specific antibodies against 16 L. interrogans serovars demonstrated by the microagglutination test.

Agglutination Tests↗