ATP synthase of E. coli: F1-ATPase activity is functionally decoupled from the proton-transporting complex (FO) by tributyltin.

The F-type ATP synthase/ATPase (FOF1) is important for cellular bioenergetics in eukaryotes and bacteria. We recently showed that venturicidins, a class of macrolides that inhibit the proton transporting complex (FO), can also induce time-dependent functional decoupling of F1-ATPase from FO on membranes from Escherichia coli and Pseudomonas aeruginosa. This dysregulated ATPase activity could deplete bacterial ATP levels and contribute to venturicidin's capacity to enhance the bactericidal action of aminoglycosides antibiotics. We now show that a distinct type of FO inhibitor, tributyltin, also can decouple FOF1-ATPase activity of E. coli membranes. In contrast to the action of venturicidins, decoupling by tributyltin is not dependent on ATP, indicating mechanistic differences. Tributyltin disrupts the coupling role of the ε subunit of F1 but does not induce dissociation of the F1-ATPase complex from membrane-embedded FO. Understanding such decoupling mechanisms could support development of novel antibacterial compounds that target dysregulation of FOF1 functions.

Tributyltin-induced oxidative stress causes developmental damage in the cardiovascular system of zebrafish (Danio rerio).

Tributyltin (TBT) can be used as an antifouling agent with anticorrosive, antiseptic and antifungal properties and is widely used in wood preservation and ship painting. However, it has recently been found that TBT can be harmful to aquatic organisms. In this study, to gain insight into the effects of TBT with respect to the development of the cardiovascular system in zebrafish embryos, zebrafish embryos were exposed to different concentrations of TBT solutions (0.2 µg/L, 1 µg/L, and 2 µg/L) at 2 h post-fertilization (hpf) TBT exposure resulted in decreased hatchability and heart rate, deformed features such as pericardial edema, yolk sac edema, and spinal curvature in zebrafish embryos, and impaired heart development. Expression of cardiac development-related genes (vmhc, myh6, nkx2.5, tbx5a, gata4, tbx2b, nppa) is dysregulated. Transgenic zebrafish Tg (fli1: EGFP) were used to explore the effects of TBT exposure on vascular development. It was found that TBT exposure could lead to impaired development of intersegmental vessels (ISVs), common cardinal vein (CCV), subintestinal vessels (SIVs) and cerebrovascular. The expression of vascular endothelial growth factor (VEGF) signaling pathway-related genes (flt1, flt4, kdr, vegfa) was downregulated. Biochemical indices showed that ROS and MDA levels were significantly elevated and that SOD and CAT activities were significantly reduced. The expression of key genes for prostacyclin synthesis (pla2, ptgs2a, ptgs2b, ptgis, ptgs1) is abnormal. Therefore, it is possible that oxidative stress induced by TBT exposure leads to the blockage of arachidonic acid (AA) production in zebrafish embryos, which affects prostacyclin synthesis and consequently the normal development of the heart and blood vessels in zebrafish embryos.

Whole-Transcriptome Analysis Reveals the RNA Profiles in Mouse Bone Marrow Mesenchymal Stem Cells or Zebrafish Embryos After Exposure to Environmental Level of Tributyltin.

To understand the underlying molecular mechanisms, mouse bone marrow mesenchymal stem cells (BMSCs) and zebrafish embryos were exposed to the control group and Tributyltin (TBT) group (10 ng/L, environmental concentration) for 48 h, respectively. The expression profiles of RNAs were investigated using whole-transcriptome analysis in mouse BMSCs or zebrafish embryos after TBT exposure. For mouse BMSCs, the results showed 2,449 differentially expressed (DE) mRNAs, 59 DE miRNAs, 317 DE lncRNAs, and 15 circRNAs. Similarly, for zebrafish embryos, the results showed 1,511 DE mRNAs, 4 DE miRNAs, 272 DE lncRNAs, and 28 circRNAs. According to KEGG pathway analysis showed that DE RNAs were mainly associated with immune responses, signaling, and cellular interactions. Competing endogenous RNA (ceRNA) network analysis revealed that the regulatory network of miRNA-circRNA constructed in zebrafish embryos was more complex compared to that of mouse BMSCs.

Low-dose tributyltin triggers human chondrocyte senescence and mouse articular cartilage aging.

Tributyltin (TBT) is known as an endocrine-disrupting chemical. This study investigated the effects and possible mechanisms of TBT exposure on inducing human articular chondrocyte senescence in vitro at the human-relevant concentrations of 0.01-0.5 µM and mouse articular cartilage aging in vivo at the doses of 5 and 25 µg/kg/day, which were 5 times lower than the established no observed adverse effect level (NOAEL) and equal to NOAEL, respectively. TBT significantly increased the senescence-associated ß-galactosidase activity and the protein expression levels of senescence markers p16, p53, and p21 in chondrocytes. TBT induced the protein phosphorylation of both p38 and JNK mitogen-activated protein kinases in which the JNK signaling was a main pathway to be involved in TBT-induced chondrocyte senescence. The phosphorylation of both ataxia-telangiectasia mutated (ATM) and histone protein H2AX (termed γH2AX) was also significantly increased in TBT-treated chondrocytes. ATM inhibitor significantly inhibited the protein expression levels of γH2AX, phosphorylated p38, phosphorylated JNK, p16, p53, and p21. TBT significantly stimulated the mRNA expression of senescence-associated secretory phenotype (SASP)-related factors, including IL-1ß, TGF-ß, TNF-α, ICAM-1, CCL2, and MMP13, and the protein expression of GATA4 and phosphorylated NF-κB-p65 in chondrocytes. Furthermore, TBT by oral gavage for 4 weeks in mice significantly enhanced the articular cartilage aging and abrasion. The protein expression of phosphorylated p38, phosphorylated JNK, GATA4, and phosphorylated NF-κB-p65, and the mRNA expression of SASP-related factors were enhanced in the mouse cartilages. These results suggest that TBT exposure can trigger human chondrocyte senescence in vitro and accelerating mouse articular cartilage aging in vivo.

Organotin and organochlorine toxicants activate key translational regulatory proteins in human immune cells.

Environmental contaminant exposures occur due to the widespread use of synthetic chemicals. Tributyltin (TBT), dibutyltin (DBT), and pentachlorophenol (PCP) are each used in a variety of applications, including antifouling paints and stabilizers in certain plastics. Each of these compounds has been found in human blood, as well as other tissues, and they have been shown to stimulate pro-inflammatory cytokine production in human immune cells, Inflammatory cytokines mediate response to injury or infection. However, if their levels are increased in the absence of an appropriate stimulus, chronic inflammation can occur. Chronic inflammation is associated with a number of pathologies including cancer. Stimulation of pro-inflammatory cytokine production by these toxicants is dependent on activation of ERK 1/2 and/or p38 MAPK pathways. MAPK pathways have the capacity to regulate translation by increasing phosphorylation of key translation regulatory proteins. There have been no previous studies examining the effects of TBT, DBT, or PCP on translation. The current study shows that ribosomal protein S6 (S6), eukaryotic initiation factor 4B (eIF4B), and eIF4E are phosphorylated (activated) and/or their total levels are elevated in response to each of these compounds at concentrations found in human blood. Activation/increased levels of translational proteins occurred at concentrations of the compounds that have been shown to elevate pro-inflammatory cytokine production, but where there is no increase in mRNA for those proteins was seen. Compound-stimulated increases in translation appear to be part of the mechanism by which they elevate protein production in immune cells.

Biochemical mechanisms of tributyltin chloride-induced cell toxicity in Sertoli cells.

Tributyltin chloride (TBTCL) is a widely used fungicide and heat stabilizer in compositions of PVC. TBTCL has been detected in human bodies and potentially causes harmful effects on humans' thyroid, cardiovascular and other organs. As one of the first examples of endocrine disruptors, the toxicity effects of TBTCL on the male reproduction system have aroused concerns. However, the potential cellular mechanisms are not fully explored. In the current study, by using Sertoli cells, a critical regulator of spermatogenesis as a cell model, we showed that with 200 nM exposure for 24 h, TBTCL causes apoptosis and cell cycle arrest. RNA sequencing analyses suggested that TBTCL probably activates endoplasmic reticulum (ER) stress, and disrupts autophagy. Biochemical analysis showed that TBTCL indeed induces ER stress and the dysregulation of autophagy. Interestingly, activation of ER stress and inhibition of autophagy is responsible for TBTCL-induced apoptosis and cell cycle arrest. Our results thus uncovered a novel insight into the cellular mechanisms for TBTCL-induced toxicology in Sertoli cells.

Embryonic exposure to environmentally relevant levels of tributyltin affects embryonic tributyltin bioaccumulation and the physiological responses of juveniles in cuttlefish (Sepia pharaonis).

Tributyltin (TBT) is a typical organic pollutant that persists in aquatic sediments due to its wide usage as an antifouling fungicide during the past few decades. Despite increased awareness of the serious negative consequences of TBT on aquatic species, studies on the effects of TBT exposure on cephalopod embryonic development and juvenile physiological performance are scarce. To investigate the lasting effects of TBT toxicity on Sepia pharaonis from embryo to hatchling, embryos (gastrula stage, 3-5 h post fertilization) were exposed to four levels of TBT until hatching: 0 (control), 30 (environmental level), 60, and 120 ng/L. Subsequently, juvenile growth performance endpoints and behavioral alterations were assessed over 15 days post-hatching. Egg hatchability was significantly reduced and embryonic development (i.e., premature hatching) was accelerated in response to 30 ng/L TBT exposure. Meanwhile, TBT-induced alterations in embryonic morphology primarily included yolk-sac lysis, embryonic malformations, and uneven pigment distributions. During the pre-middle stage of embryonic development, the eggshell serves as an effective barrier to safeguard the embryo from exposure to 30-60 ng/L TBT, according to patterns of TBT accumulation and distribution in the egg compartment. However, even environmental relevant levels of TBT (30 ng/L) exposure during embryonic development had a negative impact on juvenile behavior and growth, including slowing growth, shortening eating times, causing more irregular movements, and increasing inking times. These findings indicate that after TBT exposure, negative long-lasting effects on S. pharaonis development from embryo to hatchling persist, suggesting that long-lasting toxic effects endure from S. pharaonis embryos to hatchlings.

The possible ameliorative role of Lycopene on Tributyltin induced thyroid damage in adult male albino rats (histological, immunohistochemical and biochemical study).

Tributyltin is used in industrial applications. This current research aimed to study the effect of Tributyltin on the thyroid gland structure and function of adult male albino rats and the protective effect of Lycopene. Twenty-one male adult albino rats were classified into three groups: Control, treated that received Tributyltin, and protective that received Lycopene with Tributyltin. At the end of the experiment, blood samples were collected and T4, T3, and (TSH) were measured. Tissue superoxide dismutase (SOD) and malondialdehyde (MDA) were estimated. Thyroid gland specimens were processed for histological and immunohistochemical examination. Then morphometric and statistical analyses were done. The treated group showed affection in thyroid function and histological structure as vacuolated colloid and cytoplasm and dark nuclei. Ultrastructurally, follicular cells showed irregular shrunken nuclei, atrophied apical microvilli, vacuoles, multiple lysosomal granules, mitochondria with destructed cristae, and extensively dilated rough endoplasmic reticulum. There was increase in Caspase-3 immunoexpression and decrease in Beclin-1 immunoexpression. The thyroid structure and biochemical markers improved after Lycopene administration. The thyroid gland damage caused by Tributyltin is ameliorated by Lycopene.

Evaluating the histomorphological and biochemical changes induced by Tributyltin Chloride on pituitary-testicular axis of adult albino rats and the possible ameliorative role of hesperidin.

This study was performed to explore in detail the toxic effects of Tributyltin Chloride (TBT) on the pituitary-testicular axis and the possible amelioration with Hesperidin. Seventy-two adult male albino rats were divided into four groups: Control group (I), TBT-treated group (II), TBT+Hesperidin group (III), and Recovery group (IV). Body and testicular weights were measured. Blood samples were taken to estimate serum levels of testosterone, FSH and LH hormones by enzyme-linked immunosorbent assay (ELISA). Malondialdehyde (MDA) level was measured in testes homogenates. Tissue samples from the pituitary glands and testes were processed for light, electron microscope examination, and immunohistochemical detection of anti-FSH, and Ki67 proteins. Results showed a statistically significant decrease in testicular weight, serum testosterone, FSH and LH levels and a significant increase in tissue MDA in the TBT group when compared to the control group. TBT treatment caused severe histopathological changes with decreased area percent of PAS-stained basophils, and anti FSH immuno-stained gonadotrophs in the pituitary gland. The testes of group II also showed marked tissue damage, cell loss with decreased epithelial height and decreased number of proliferating spermatogenic cells. Hesperidin supplementation with TBT proved significant amelioration of the previously mentioned parameters in both glands which could improve male fertility. In conclusion: The flavonoid Hesperidin has the potential to protect against the reproductive damage induced by TBT in susceptible individuals.

Retinoids promote penis development in sequentially hermaphroditic snails.

Sexual systems are surprisingly diverse, considering the ubiquity of sexual reproduction. Sequential hermaphroditism, the ability of an individual to change sex, has emerged multiple times independently across the animal kingdom. In molluscs, repeated shifts between ancestrally separate sexes and hermaphroditism are generally found at the level of family and above, suggesting recruitment of deeply conserved mechanisms. Despite this, molecular mechanisms of sexual development are poorly known. In molluscs with separate sexes, endocrine disrupting toxins bind the retinoid X receptor (RXR), activating ectopic male development in females, suggesting the retinoid pathway as a candidate controlling sexual transitions in sequential hermaphrodites. We therefore tested the role of retinoic acid signaling in sequentially hermaphroditic Crepidula snails, which develop first into males, then change sex, maturing into females. We show that retinoid agonists induce precocious penis growth in juveniles and superimposition of male development in females. Combining RXR antagonists with retinoid agonists significantly reduces penis length in induced juveniles, while similar treatments using retinoic acid receptor (RAR) antagonists increase penis length. Transcripts of both receptors are expressed in the induced penis. Our findings therefore show that retinoid signaling can initiate molluscan male genital development, and regulate penis length. Further, we show that retinoids induce ectopic male development in multiple Crepidula species. Species-specific influence of conspecific induction of sexual transitions correlates with responsiveness to retinoids. We propose that retinoid signaling plays a conserved role in molluscan male development, and that shifts in the timing of retinoid signaling may have been important for the origins of sequential hermaphroditism within molluscs.