Prednisone and prednisolone effects on development, blood, biochemical and histopathological markers of Aquarana catesbeianus tadpoles.

Synthetic glucocorticoids are often found in surface waters and can cause harmful effects to aquatic organisms such as amphibians. In this work we evaluated the effects of the drugs prednisone (PD) and prednisolone (PL) on developmental, molecular, blood, biochemical and histological markers. Aquarana catesbeianus tadpoles were exposed for 16 days to environmentally relevant concentrations of 0, 0.1, 1 and 10 µg/L of both drugs. PD increased the transcript levels of the enzyme deiodinase III (Dio3), the hormones cortisol and T4 and delayed development. Changes in the thyroid gland occurred after tadpoles were exposed to both drugs, with a reduction in the diameter and number of follicles and an increase/or decrease in area. Also, both drugs caused a decrease in lymphocytes (L) and an increase in neutrophils (N), thrombocytes, the N:L ratio and lobed and notched erythrocytes. Increased activity of the enzymes superoxide dismutase, glutathione S-transferase and glucose 6-phosphate dehydrogenase was observed after exposure to PD. Furthermore, both drugs caused an increase in the activity of the enzymes catalase and glutathione peroxidase. However, only PD caused oxidative stress in exposed tadpoles, evidenced by increased levels of malondialdehyde and carbonyl proteins. Both drugs caused an increase in inflammatory infiltrates, blood cells and melanomacrophages in the liver. Our results indicate that PD was more toxic than PL, affecting development and causing oxidative stress.

Prednisolone Accelerates Embryonic Development of Zebrafish via Glucocorticoid Receptor Signaling at Low Concentrations.

Synthetic glucocorticoids have been widely detected in aquatic ecosystems and may pose a toxicological risk to fish. In the present study, we described multiple end point responses of zebrafish to a commonly prescribed glucocorticoid, prednisolone (PREL), at concentrations between 0.001 and 9.26 µg/L. Of 23 end points monitored, 7 were affected significantly. Significant increases in the frequency of yolk extension formation, spontaneous contraction, heart rate, and ocular melanin density and significant decreases of ear-eye distance at PREL concentrations of 0.001 µg/L and above clearly pointed to the acceleration of embryonic development of zebrafish by PREL. Further confirmation came from the alterations in somite numbers, head-trunk angle, and yolk sac size, as well as outcomes obtained via RNA sequencing, in which signaling pathways involved in tissue/organ growth and development were highly enriched in embryos upon PREL exposure. In addition, the crucial role of glucocorticoid receptor (GR) for PREL-induced effects was confirmed by both, the coexposure to antagonist mifepristone (RU486) and GR-/- mutant zebrafish experiments. We further demonstrated similar accelerations of embryonic development of zebrafish upon exposure to 11 additional glucocorticoids, indicating generic adverse effect characteristics. Overall, our results revealed developmental alterations of PREL in fish embryos at low concentrations and thus provided novel insights into the understanding of the potential environmental risks of glucocorticoids.

In vitro study of the toxic and teratogenic effects of prednisolone, azathioprine and mycophenolate mofetile on embryological development of rats.

This study aimed to evaluate the effects of the glucocorticoid prednisolone, the mycophenolic acid prodrug, azathioprine, and the fungi fermentation end product, mycophenolate mofetile on the embryological development of rats. Nine day-old rat embryos were cultured in rat serum containing prednisolone at varying concentrations (5-30 µg/ml) for 48 h. The test groups were cultured separately in rat serum containing 0.3-10 µg/ml azathioprine and 1-10 µg/ml mycophenolate mofetile. Embryonic development parameter effects of both drugs in combination with prednisolone (20 µg/ml) were studied using morphological methods, with special attention given to the incidence of malformations. The genotoxic effects of agents evaluated with the TUNEL test revealed that prednisolone is not a cause of developmental toxicity. The maximum safe dose of prednisolone that could be used in combination with other immunosuppressive agents was determined to be 20 µg/ml. Azathioprine was found to be toxic and teratogenic for the rat embryos beginning at a dose of 1 µg/ml. Dose-dependent toxic and teratogenic effects of mycophenolate mofetile were detected at doses lower than normal clinical ones.

Three dimensional and microphysiological bone marrow models detect in vivo positive compounds.

Micronucleus (MN) assessment is a valuable tool in safety assessment. However, several compounds are positive in the in vivo bone marrow (BM) MN assay but negative in vitro, reflecting that BM complexity is not recapitulated in vitro. Importantly, these compounds are not genotoxic; rather, drug-driven pharmacological-effects on the BM increase MN, however, without mechanistic understanding, in vivo positives stop drug-progression. Thus, physiologically-relevant BM models are required to bridge the gap between in vitro and in vivo. The current study aimed to investigate the utility of two human 3D BM models (fluidic and static) for MN assessment. MN induction following treatment with etoposide and Poly-ADP Ribose Polymerase inhibitor (PARPi) and prednisolone (negative in vitro, positive in vivo) was determined in 2D L5178Y and human BM cells, and the 3D BM models. Etoposide (0-0.070 µM) and PARPi (0-150 µM) induced MN in both 3D BM models indicating their utility for genotoxicity testing. Interestingly, PARPi treatment induced a MN trend in 3D more comparable to in vivo. Importantly, prednisolone (0-1.7 mM) induced MN in both 3D BM models, suggesting recapitulation of the in vivo microenvironment. These models could provide a valuable tool to follow up, and eventually predict, suspected pharmacological mechanisms, thereby reducing animal studies.

Prednisolone suppresses collagen-encoding gene expression causing cartilage defects in zebrafish larvae.

Glucocorticoid-induced osteoporosis (GIOP) is a clinically important disease. Despite many studies, the intrinsic pathogenesis of GIOP is still not fully understood. Cartilage is the target tissue of the glucocorticoid prednisolone (PN). To explore the intrinsic mechanism of PN-induced cartilage damage, we performed cartilage staining and cell transfection experiments in zebrafish larvae treated with PN. The results showed that PN caused cartilage damage in zebrafish at 25 µM. Moreover, after treatment with PN, it was found that collagen-encoding gene expression was significantly reduced. Further research revealed that the glucocorticoid receptor (GR) mediates the transcriptional inhibition of collagen genes by PN. These results indicate that glucocorticoids cause cartilage damage by inhibiting the expression of collagen genes through their receptors. Our study provides new insights into GIOP.

Effects of prednisolone on behavior and hypothalamic-pituitary-interrenal axis activity in zebrafish.

Prednisolone is a synthetic glucocorticoid used clinically for treating allergies, inflammation, and autoimmune diseases. Long-term prednisolone use has been shown to have negative effects on physiology and mood. We aimed to study the pharmacology and toxicology of glucocorticoid-like drugs by investigating behavioral and hypothalamic-pituitary-interrenal (HPI) axis effects in a zebrafish model. Zebrafish embryos 24 h post fertilization were exposed to 25 µM prednisolone. Their behavior was investigated 5 days post fertilization (dpf), and their HPI axis-related activity and related neurotransmitter levels were investigated 3, 4, 5, and 6 dpf. The behavior results showed that exposure to prednisolone resulted in decreased autonomic activity and low sensitivity to light. qRT-PCR and ELISA results showed decreased activity of the HPI axis and increased secretion of dopamine and serotonin after exposure to prednisolone. This study provides us with new insights into understanding the effects of glucocorticoids on the HPI axis.

Vitamin E and Selenium Reduce Prednisolone Side Effects in Rat Hearts.

Aim of this work was to determine the effects of dietary intake vitamin E and Se on lipid peroxidation (LPO) as Thiobarbituric acid reactive substances (TBARS) and on the antioxidative defense mechanisms in heart tissues of rats treated with high doses of prednisolone. 250 adult male Wistar rats were randomly divided into 5 groups and fed with normal diet. Additionally groups 3, 4, and 5 received a daily supplement in their drinking water of 20 mg vitamin E, 0.3 mg Se, and a combination of vitamin E and Se (20 mg/ 0.3 mg), respectively, for 30 days. For 3 d subsequently, control group was treated with placebo, and remaining four groups were injected intramuscularly with 100 mg/kg prednisolone. After last administration of prednisolone, 10 rats from each group were killed at 4, 8, 12, 24, and 48 h and the activities of antioxidant enzymes and the levels of GSH and TBARS were measured. GSH-Px, CAT activities and GSH levels decreased starting from 4th hour to 48% and 65% of control levels by 24th hour, respectively and it reincreased to control levels at 48th hour in the prednisolone group (p < 0.001, p < 0.001). In addition, prednisolone administration led 2-fold increase in heart TBARS levels at 24th hour (p < 0.001). E vitamins and Se inhibited the increase in heart TBARS and the decrease in antioxidative enzymes levels. Therefore, It is concluded that vitamin E and Se may have a preventive role in decreasing the increase of TBARS caused by prednisolone administration in our study.

Treatment with synthetic glucocorticoid impairs bone metabolism, as revealed by in vivo imaging of osteoblasts and osteoclasts in medaka fish.

Glucocorticoids (GCs) play an important role in the stress reaction and function in the development of multiple tissues in our body. When given chronically in supraphysiologic doses, GCs are associated with orthodontic tooth movement, with serious side effects and particularly adverse effects on bone metabolism. However, the effects of steroids on bone cell dynamics are incompletely understood. Therefore, in this present study we examined the participation of osteoblasts and osteoclasts in osterix-DsRed/TRAP-EGFP double transgenic (Tg) medaka treated with synthetic GCs. Chronic continuous administration of prednisolone (PN) significantly reduced the fluorescence signals in the whole body of both osterix-DsRed and TRAP-EGFP medaka at 18 days, and those of the pharyngeal bone and tooth region at 32 days. To examine the capacity of the medaka for fracture healing during chronic administration of PN, we caused a fracture of a part of the bony fin ray at 18 days after the initiation of PN continuous administration. The bone fracture healing was significantly delayed by 32 days, accompanied by decreased signal area of both osterix-DsRed and TRAP-EGFP compared with that of the control. Next, to investigate the effect of acute administration of GC on the fracture healing, we started administration of dexamethasone (DX) immediately after the bone fracture, and this administration lasted during the 11 days of fracture healing. The results showed that the TRAP-EGFP-positive osteoclasts were reduced in area, but not the osterix-DsRed-positive osteoblasts. Lastly, to confirm the function of the glucocorticoid receptor in bone healing, we generated glucocorticoid receptor 2-deficient medaka (gr2-/-). The fluorescent signal area of osterix-DsRed and TRAP-EGFP were increased at bone fracture sites in these fish, and DX treatment of them decreased the TRAP-EGFP signal area compared with that for the control fish. Our results indicate that GRs negatively regulated osteoclast recruitment and the accumulation of osteoblasts in bone fracture healing.

Toxicology study for magnetic injection of prednisolone into the rat cochlea.

This paper investigates the safety of a novel 'magnetic injection' method of delivering therapy to the cochlea, in a rodent model. In this method of administration, a magnetic field is employed to actively transport drug-eluting superparamagnetic iron-oxide core nanoparticles into the cochlea, where they then release their drug payload (we delivered the steroid prednisolone). Our study design and selection of control groups was based on published regulatory guidance for safety studies that involve local drug delivery. We tested for both single and multiple delivery doses to the cochlea, and found that magnetic delivery did not harm hearing. There was no statistical difference in hearing between magnetically treated ears versus ears that received intra-tympanic steroid (a mimic of a standard-of-care for sudden sensorineural hearing loss), both 2 and 30 days after treatment. Since our treatment is local to the ear, the levels of steroid and iron circulating systemically after our treatment were low, below mass-spectrometry detection limits for the steroid and no different from normal for iron. No adverse findings were observed in ear tissue histopathology or in animal gross behavior. At 2 and 30 days after treatment, inflammatory changes examined in the ear were limited to the middle ear, were very mild in severity, and by day 90 there was ongoing and almost complete reversibility of these changes. There were no ear tissue scarring or hemorrhage trends associated with magnetic delivery. In summary, after conducting a pre-clinical safety study, no adverse safety issues were observed.

Prednisolone degradation by UV/chlorine process: Influence factors, transformation products and mechanism.

Prednisolone (PDNN) as an emergent micropollutant directly influences the regional ecological security. In this study, the degradation of PDNN by ultraviolet activated chlorine (UV/chlorine) oxidation process was comprehensively evaluated. The quenching experiment suggested that the PDNN degradation in UV/chlorine process was involved in the participation of hydroxyl radical (OH) and reactive chlorine species (RCS). Influence factors including chlorine dosage, pH, common anion and cation, fulvic acid (FA) on PDNN degradation via UV/chlorine process were investigated. A low chlorine (≤7.1 mg L-1) promoted the PDNN degradation, while a high chlorine dosage (>7.1 mg L-1) was adverse. The pH (4.0-10.0) showed negligible effect, while the investigated anions (Cl-, Br-, HCO3- and SO42-), NH4+ and FA exerted negative impact on PDNN degradation. An efficient process to minimize pharmaceutical micropollutants was the disposal of human urine containing a high concentration of pharmaceutical and potential toxic metabolites. An inhibitory effect was observed in the synthetic urine (fresh urine and hydrolyzed urine). The intermediates/products were identified and the mechanism of PDNN degradation was proposed. PDNN gone through three degradation routes, involving the direct addition of α, ß-unsaturated ketone at C1 or C5, the photolysis of C17 and H-abstraction of C11. The main reactive sites were further determined by comparison of the frontier orbitals calculation and the proposed mechanism. Based on the toxicological tests for PDNN degradation, TP396 (TP396-C1Cl and TP396-C5Cl) and TP414-2-1 (TP414-C1ClC5OH) exhibited much higher toxicity than PDNN, and prolonging reaction time was necessary to achieve PDNN detoxification.

Cytotoxicity of anticancer candidate salinomycin and identification of its metabolites in rat cell cultures.

Salinomycin (SAL) is a polyether antibiotic, which is commonly used as a coccidiostat and has recently shown to exhibit anticancer activity. The toxic action of the drug may be connected with the extent and routes of its biotransformation. The cytotoxic potential of SAL and its combination with tiamulin and prednisolone was investigated using three cell models from rat: primary hepatocytes, hepatoma cells (FaO) and myoblasts (L6). The four biochemical endpoints were assessed: mitochondrial and lysosomal activity, total cell protein content and membrane integrity. The metabolites of SAL in the medium from cell cultures were determined using LC-MS/MS. The cytotoxicity of SAL was time-, concentration- and cells dependent. The most sensitive endpoint was the inhibition of lysosomal activity. Tiamulin increased SAL cytotoxicity, whereas the opposite results were observed for prednisolone. Primary hepatocytes were the most efficient in SAL biotransformation both in terms of its intensity and number of produced metabolites. The range of the cytotoxicity and mode of salinomycin interaction with tiamulin and prednisolone cannot be explained by the biotransformation alone.

Environmental impact of switching from the synthetic glucocorticoid prednisolone to the natural alkaloid berberine.

Low amounts of human pharmaceuticals in the aquatic environment can affect bacteria, animals and ultimately humans. Here, the environmental consequences of a shift in prescription behavior from prednisolone to berberine was modeled using an environmental decision support system based on four consecutive steps: emission, fate, exposure and effect. This model estimates the relative aquatic and human health impacts of alternative pharmaceutical prescriptions throughout Europe. Since a Defined Daily Dose (DDD) of berberine has yet to be formulated, the environmental impacts of berberine and prednisolone were compared under the assumption of equal DDDs. Subsequently, the relative impact ratio indicates the extent to which the actual DDD of berberine might be higher to still be environmentally preferable over prednisolone. In fact, berberine can be administered at a six times higher dose throughout Europe before its impact on the aquatic environment exceeds that of one prescription of prednisolone. On average, the results for impacts on human health are similar, with the median impact ratio ranging between 5.87 and 22.8 depending on the level of drinking water purification. However, for some regions in Spain, Austria, Baltic States and Finland, berberine can only be considered an environmentally better alternative if it is administered at a lower dose than prednisolone. We conclude that for most regions in Europe it is, up until a certain dose of berberine, beneficial for the aquatic environment and therefore human health to prefer prescription of berberine over prednisolone.

Nano-engineering of biomedical prednisolone liposomes: evaluation of the cytotoxic effect on human colon carcinoma cell lines.

OBJECTIVES: Liposomes have attracted the attention of researchers due to their potential to act as drug delivery systems for cancer treatment. The present investigation aimed to develop liposomes loaded with prednisolone base and the evaluation of the antiproliferative effect on human colon carcinoma cell lines. METHODS: Liposomes were elaborated by following a reproducible thin film hydration technique. The physicochemical characterization of liposomes included photon correlation spectroscopy, microscopy analysis, Fourier transform infrared spectroscopy, rheological behaviour and electrophoresis. On the basis of these data and drug loading values, the best formulation was selected. Stability and drug release properties were also tested. KEY FINDINGS: Resulting liposomes exhibited optimal physicochemical and stability properties, an excellent haemocompatibility and direct antiproliferative effect on human colon carcinoma T-84 cell lines. CONCLUSIONS: This study shows direct antitumour effect of prednisolone liposomal formulation, which opens the door for liposomal glucocorticoids as novel antitumour agents.

Assessing multigenerational effects of prednisolone to the freshwater snail, Physa acuta (Gastropoda: Physidae).

Prednisolone (PDS), a potent synthetic glucocorticoid is widely prescribed for its exceptional anti-inflammatory properties. Several studies have detected the environmental presence of PDS in water bodies which has led to an ecological concern for its toxicity to non-target aquatic biota. The present study investigated the effects of exposure to PDS on different life-cycle stages and generations of the freshwater snail, Physa acuta. This continuous exposure over a period of multiple generations resulted in generational impairments at measured endpoints. LOEC values (p<0.001) for PDS exposure ranged from 32 to 4µg/L in exposed F0-F2 generations. Global DNA methylation (% 5-methyl cytosine) of adult progeny was found to be affected at higher test concentrations in comparison to the parent snails. Partially formed to completely missed growth components of shell structure and shell thinning in abnormally underdeveloped PDS exposed snails of F1 and F2 generation, was also observed in this multigenerational exposure experiment. The multigenerational study confirmed P. acuta as a promising bioindicator since critical effects of the long term glucocorticoid exposure opens up the way for further investigations on transgenerational toxicity in environmental toxicology and risk assessment and to monitor glucocorticoid pollution in aqueous ecosystem.

Multigenerational effects of two glucocorticoids (prednisolone and dexamethasone) on life-history parameters of crustacean Ceriodaphnia dubia (Cladocera).

Synthetic glucocorticoids (GCs) such as dexamethasone (DEX) and prednisolone (PDS) have been used since the 1940s to cure inflammatory and auto-immune disorders. Their use has been linked to a host of deleterious effects in aquatic ecosystems such as osteoporosis in vertebrates, developmental impairments in molluscs and reduced fecundity and growth in cladocerans. Apart from these handful of studies, the effects of GCs on aquatic biota are largely unknown. The present study is a first of its kind aiming to assess the multi-generational exposure effects of DEX and PDS on the life history parameters of Ceriodaphnia dubia (C. dubia). Multigenerational studies have proved to be an advantage in assessing the cumulative damage caused by aquatic toxicants at the population level of the exposed organisms over a period of successive generations using multiple biological endpoints. Test results demonstrated that C. dubia exhibited varied sensitivities towards both the studied chemicals however were more sensitive to DEX with 48-h EC50 (95% confidence interval) of 0.75 mg/L (CI: 0.59-0.92) in comparison to PDS [19 mg/L (CI: 15-23)]. EC10 values for F0 in a multigenerational chronic bioassays were 48 µg/L (CI: 37.4-61) for DEX and 460 µg/L (CI: 341-606) for PDS and in F3 were 2.2 µg/L (CI: 1.6-3.1) for DEX and 31 µg/L (CI: 19.4-46) for PDS. There was a positive trend of increased toxicity followed by reduced life history traits such as fecundity, brood size and time to first brood and intrinsic rate of population increase and body growth (length and area) of C. dubia in the case of both studied chemicals. The results from the current work highlighted the importance of multigenerational studies in identifying the evolutionary responses of stressed non-target aquatic organisms, and data obtained can be further used in developing water quality guidelines.

Vitamin D3 protects against prednisolone-induced liver injury associated with the impairment of the hepatic NF-κB/iNOS/NO pathway.

The study was carried out to define whether prednisolone-induced damage to hepatic cells is accompanied by excessive nitric oxide (NO) levels associated with nuclear factor kappa B (NF-κB)/inducible NO synthase (iNOS) activation and evaluate the efficacy of the treatment with vitamin D3. Histopathological examination, activities of liver transaminases (alanine aminotransferase and aspartate aminotransferase), and cell death assays consistently showed that prednisolone (5 mg/kg body weight, 30 days) induces chronic liver injury in female Wistar rats. Specifically, increased hepatocellular necrosis and caspase-3-dependent apoptosis were observed. Prednisolone enhanced iNOS protein expression, NO generation, and tyrosine nitration in liver cells. Despite unchanged hepatic level of the NF-κB/p65 protein, prednisolone increased inhibitory κB-α (IκB-α) degradation, nuclear translocation, and phosphorylation of NF-κB/p65 at Ser311, indicating that NF-κB activation can be involved in the induction of iNOS/NO. All changes were associated with a 2.9-fold decrease in the serum content of 25-hydroxyvitamin D3 and significant reduction of hepatic vitamin D3 receptor (VDR) expression that points reliably to vitamin D3 deficiency and failures in VDR signaling. Vitamin D3 co-administration (100 IU/rat, 30 days) prevented glucocorticoid-evoked abnormalities in hepatic tissue. In conclusion, prednisolone-induced liver disturbances were associated with the impairment of NF-κB/iNOS/NO responses that can be ameliorated by vitamin D3 treatment through VDR-mediated mechanisms.

Immunosuppressive Treatment Alters Secretion of Ileal Antimicrobial Peptides and Gut Microbiota, and Favors Subsequent Colonization by Uropathogenic Escherichia coli.

BACKGROUND: Transplant recipients are treated with immunosuppressive (IS) therapies, which impact host-microbial interactions. We examined the impact of IS drugs on gut microbiota and on the expression of ileal antimicrobial peptides. METHODS: Mice were treated for 14 days with prednisolone, mycophenolate mofetil, tacrolimus, a combination of these 3 drugs, everolimus, or water. Feces were collected before and after treatment initiation. Ileal samples were collected after sacrifice. Fecal and ileal microbiota were analyzed by pyrosequencing of 16S rRNA genes and enumeration of selected bacteria by culture, and C-type lectins were assessed in ileal tissues by reverse transcriptase-quantitative polymerase chain reaction. RESULTS: Prednisolone disrupted fecal microbiota community structure, decreased Bacteroidetes, and increased Firmicutes in the feces. Prednisolone, tacrolimus, and mycophenolate mofetil modified fecal microbiota at the family level in each experimental replicate, but changes were not consistent between the replicates. In ileal samples, the genus Clostridium sensu stricto was dramatically reduced in the prednisolone and combined IS drug groups. These modifications corresponded to an altered ileal expression of C-type lectins Reg3γ and Reg3ß, and of interleukin 22. Interestingly, the combined IS treatment enabled a commensal Escherichia coli to flourish, and dramatically increased colonization by uropathogenic E. coli strain 536. CONCLUSIONS: IS treatment alters innate antimicrobial defenses and disrupts the gut microbiota, which leads to overgrowth of indigenous E. coli and facilitates colonization by opportunistic pathogens.

Environmental concentrations of prednisolone alter visually mediated responses during early life stages of zebrafish (Danio rerio).

The development of the eye in vertebrates is dependent upon glucocorticoid signalling, however, specific components of the eye are sensitive to synthetic glucocorticoids. The presence of synthetic glucocorticoids within the aquatic environment may therefore have important consequences for fish, which are heavily reliant upon vision for mediating several key behaviours. The potential ethological impact of synthetic glucocorticoid oculotoxicity however has yet to be studied. Physiological and behavioural responses which are dependent upon vision were selected to investigate the possible toxicity of prednisolone, a commonly occurring synthetic glucocorticoid within the environment, during early life stages of zebrafish. Although exposure to prednisolone did not alter the morphology of the external eye, aggregation of melanin within the skin in response to increasing light levels was impeded and embryos exposed to prednisolone (10 µg/l) maintained a darkened phenotype. Exposure to prednisolone also increased the preference of embryos for a dark environment within a light dark box test in a concentration dependent manner. However the ability of embryos to detect motion appeared unaffected by prednisolone. Therefore, while significant effects were detected in several processes mediated by vision, changes occurred in a manner which suggest that vision was in itself unaffected by prednisolone. Neurological and endocrinological changes during early ontogeny are considered as likely candidates for future investigation.

Investigation of developmental toxicity and teratogenicity of cyclosporine A, tacrolimus and their combinations with prednisolone.

In this study, it was aimed to investigate the toxic and teratogenic effects of cyclosporine A and tacrolimus and their combinations with prednisolone using an in vitro rat embryo culture technique. Cyclosporine A (4-40 µg/ml), tacrolimus (1-20 µg/ml) and combinations of these drugs with prednisolone (20 µg/ml) at different concentrations were tested. Cyclosporine A and its combination with prednisolone were determined to have toxic effects on embryonic growth after 10 µg/ml. When used alone, the lowest dose of tacrolimus had embryotoxic effects on the total morphological score and number of somites. It was determined that cyclosporine A caused hematoma at 4 µg/ml and higher doses, and tacrolimus especially at 20 µg/ml caused an open neural tube beside hematoma. It was observed that cyclosporine A at 40 µg/ml dose initiated apoptotic effects at a very low rate, prednisolone increased this effect, tacrolimus led to excessive apoptosis after 15 µg/ml, and this effect did not change with prednisolone supplement. We are of the opinion that the doses should be determined carefully when cyclosporine A and tacrolimus are required to be administered to pregnant women with prednisolone combination, as prednisolone increases the toxic effects of cyclosporine A, and increases teratogenic effects of tacrolimus.

Prednisolone impairs embryonic and posthatching development and shell formation of the freshwater snail, Physa acuta.

The aim of the present study was to investigate the lethal and sublethal effects of prednisolone exposure on the embryonic and posthatching stage of the freshwater snail, Physa acuta. The egg masses were exposed for 14 d to prednisolone concentrations ranging from 15.6 µg/L to 1000 µg/L. Treatment with prednisolone at 125 µg/L to 1000 µg/L resulted in significant decline in growth, survival, and heart rate, as well as notable abnormalities in embryonic development. Premature embryonic hatching was observed at lower concentrations of 31.25 µg/L and 62.5 µg/L, whereas delayed hatching was seen at concentrations from 125 µg/L to 1000 µg/L. To assess impacts of prednisolone exposure on the hatched juveniles, the drug exposure was extended for another 28 d. Impairment of shell development was noted in juveniles exposed to concentrations from 62.5 µg/L to 1000 µg/L at the end of 42 d, which resulted in thin and fragile shells. The thickness of shells in snails exposed to 1000 µg/L was significantly lower in comparison to those in the 15.6-µg/L and control treatments. In addition, lower calcium concentration in shells of the exposed juvenile snails at treatments of 62.5 µg/L to 1000 µg/L consequently reduced their growth. The present study confirms that continuous exposure to prednisolone can result in deleterious effects on calcium deposition, resulting in shell thinning in the freshwater snail P. acuta. Environ Toxicol Chem 2016;35:2339-2348. © 2016 SETAC.