Spatio-temporal expression patterns of glycine-rich beta proteins and cysteine-rich beta proteins in setae development of Gekko japonicus.

BACKGROUND: Setae on the pad lamellae of the Japanese gecko Gekko japonicus (Schlegel, 1836), a vital epidermal derivative, are primarily composed of cornified beta-proteins (CBPs) and play a pivotal role in adhesion and climbing. The amino acid composition of CBPs might be a determining factor influencing their functional properties. However, the molecular mechanisms governed by CBP genes with diverse amino acid compositions in setae development remain unexplored. RESULTS: Based on RNA-seq analyses, this study confirmed that all G. japonicus CBPs (GjCBPs) are involved in setae formation. Cysteine-rich CBPs encoding genes (ge-cprp-17 to ge-cprp-26) and glycine-rich CBPs encoding genes (ge-gprp-17 to ge-gprp-22) were haphazardly selected, with quantitative real-time PCR revealing their expression patterns in embryonic pad lamellae and dorsal epidermis. It is inferred that glycine-rich CBPs are integral to the formation of both dorsal scales and lamellar setae, cysteine-rich CBPs are primarily associated with setae development. Additionally, fluorescence in situ hybridization revealed spatiotemporal differences in the expression of a glycine-rich CBP encoding gene (ge-gprp-19) and a cysteine-rich CBP encoding gene (ge-cprp-17) during dorsal scales and/or lamellar development. CONCLUSIONS: All 66 CBPs are involved in the formation of setae. Glycine-rich CBPs hold a significant role in the development of dorsal scales and lamellar setae, whereas most cysteine-rich CBPs appear to be essential components of G. japonicus setae. Even GjCBPs with similar amino acid compositions may play diverse functions. The clear spatio-temporal expression differences between the glycine-rich and cysteine-rich CBP encoding genes during epidermal scale and/or setae formation were observed. Embryonic developmental stages 39 to 42 emerged as crucial phases for setae development. These findings lay the groundwork for deeper investigation into the function of GjCBPs in the development of G. japonicus setae.

Hepatic lipid accumulation is associated with multiple metabolic pathway alterations but not dyslipidemia and insulin resistance in central bearded dragons (Pogona vitticeps).

OBJECTIVE: To investigate associations between hepatic fat accumulation, fibrosis, and plasma values of primary metabolites, biochemical measurands, insulin, and lipoproteins in bearded dragons. ANIMALS: 48 adult central bearded dragons (Pogona vitticeps). METHODS: Dragons were sedated with alfaxalone, and a blood sample was collected. Plasma was submitted for untargeted primary metabolomics using gas chromatography time-of-flight mass spectrometry, a biochemistry panel, and a lipoprotein panel determined by PAGE. Hepatic lipid content was quantified by liver attenuation measurements from CT images and digital image analysis of standardized histologic sections of the liver. Fibrosis was quantified by digital image analysis on Masson's trichrome-stained histologic sections. Severity was determined from pathologic review of liver sections according to a standardized grading system. Statistical associations were investigated using serial linear models adjusted for false discovery rate and multivariate statistics. RESULTS: Both hepatic fat and fibrosis had a significant effect on CT liver attenuation values. Several oligosaccharides (maltotriose, maltose, ribose, trehalose) and alkaline phosphatase were significantly and linearly increased with hepatic lipid content (all q < .05). On partial least square-discriminant analysis, ß-hydroxybutyric acid was the most important discriminatory variable between fatty liver severity grades on histology. No significant associations were found with insulin, lipoproteins, and succinic acid. CLINICAL RELEVANCE: Bearded dragons with hepatic lipid accumulation experienced multiple metabolic pathway disruptions, some being compatible with mitochondrial dysfunction. No evidence of insulin resistance or dyslipidemia was found. Hepatic biopsy and histopathology remain recommended for reliably diagnosing and staging fatty liver disease in bearded dragons.

Pesticides compromise health: a comparison between lizards collected within and outside an agricultural area.

Reptiles are the least studied vertebrates regarding the impact of pesticides on their health, despite being good models for ecotoxicological studies given their abundance and easy handling. Salvator merianae is widely distributed in South America and often found in agricultural cultivation areas. Here, we compared the morphological, biochemical, and physiological parameters of S. merianae from an exposed area (EA) to pesticides and a reference area (RA) or control. These parameters were measured in plasma (albumin, alanine transaminase, alkaline phosphatase, gamma-glutamyl transpeptidase, glucose, total proteins, uric acid, triglycerides, VLDL, and corticosterone) and in erythrocytes (TBARS, glutathione S-transferase, superoxide dismutase, and catalase activity). Blood samples were collected from 28 lizards (EA: three juveniles, three adult females, and three adult males; RA: nine juveniles, four females, and five males) in southern Brazil during the reproductive period. We observed a decrease in body mass, the ratio between body mass and total length and snout-vent length in juvenile lizards collected at EA. The levels of TBARS, glutathione S-transferase, triglycerides, VLDL, and uric acid were altered for juveniles in EA. When comparing the two areas, females differed in superoxide dismutase activity and total proteins, while males differed in superoxide dismutase, catalase, and glutathione S-transferase activity. This set of results shows that S. merianae, especially juveniles, suffers a negative impact when inserted in an agricultural area. The analyzed biomarkers proved suitable for monitoring these lizards and the quality of this environment.

Sexually dimorphic expression of AMH facilitates testis differentiation pathway in the tropical lizard, Calotes versicolor (Daud.).

The Anti-mullerian hormone (AMH), also known as Mullerian inhibiting substance (MIS), is a glycoprotein that belongs to transforming growth factor ß superfamily. The significance of AMH during gonadal differentiation is not clearly deciphered in reptiles. Hence, current study aims to know the onset of AMH secretion and its functional role in Mullerian duct regression gonadal differentiation in tropical lizard, Calotes versicolor which exhibits a novel Female-Male-Female-Male (FMFM) pattern of temperature-dependent sex determination (TSD). The Immunohistochemistry and qRT-PCR techniques were employed to analyze the gonadal expression profile of AMH during different stages of embryonic development. The eggs of the lizard were incubated at both male-producing temperature (MPT: 25.5 ± 0.5 °C) and female-producing temperatures (FPT: 31.5 ± 0.5 °C). The results reveal that the onset of AMH gene expression was observed as early as oviposition prior to the immunolocalization of AMH protein at early-TSP (Temperature-sensitive period). The substantial rise in the intensity of the immunoreaction of AMH protein in the cytoplasm confining to Sertoli cells of seminiferous cords at MPT with low level of expression at FPT during gonadal sex differentiation, specify sexually dimorphic expression of AMH protein. Further, with the onset of sexual differentiation, the developing testis immensely expresses AMH gene which is 7-fold greater than that of transcripts levels in female embryos; signifies its conserved role in Mullerian duct regression thereby promoting testis differentiation. The robust immunnoexpression of AMH protein during post-gonadal differentiation coincides with the onset of the regression of Mullerian duct point out a positive correlation between testis differentiation and Mullerian duct regression, thus facilitating testis differentiation pathway. Based on the immunoexpression pattern of AMH protein and transcript levels of AMH gene, it is inferred that AMH plays a significant role in Mullerian duct regression, favoring testis differentiation.

Cell-permeable chameleonic peptides: Exploiting conformational dynamics in de novo cyclic peptide design.

Membrane-traversing peptides offer opportunities for targeting intracellular proteins and oral delivery. Despite progress in understanding the mechanisms underlying membrane traversal in natural cell-permeable peptides, there are still several challenges to designing membrane-traversing peptides with diverse shapes and sizes. Conformational flexibility appears to be a key determinant of membrane permeability of large macrocycles. We review recent developments in the design and validation of chameleonic cyclic peptides, which can switch between alternative conformations to enable improved permeability through cell membranes, while still maintaining reasonable solubility and exposed polar functional groups for target protein binding. Finally, we discuss the principles, strategies, and practical considerations for rational design, discovery, and validation of permeable chameleonic peptides.

Sex steroid receptors in the ovarian follicles of the lizard Sceloporus torquatus.

Estradiol and progesterone have been recognized as important mediators of reproductive events in the female mainly via binding to their receptors. This study aimed to characterize the immunolocalization of the estrogen receptor alfa (ERα), estrogen receptor beta (ERß) and progesterone receptor (PR) in the ovarian follicles of the lizard Sceloporus torquatus. The localization of steroid receptors has a spatio-temporal pattern that depends on the stage of follicular development. The immunostaining intensity of the three receptors was high in the pyriform cells and the cortex of the oocyte of previtellogenic follicles. During the vitellogenic phase, the granulosa and theca immunostaining was intense even with the modification of the follicular layer. In the preovulatory follicles, the receptors were found in yolk and additionally, ERα was also located in the theca. These observations suggest a role for sex steroids in regulating follicular development in lizards, like other vertebrates.

Individual variation within wild populations of an arid-zone lizard dictates oxidative stress levels despite exposure to sublethal pesticides.

The relationship between sublethal pesticide exposure and oxidative stress in an ecologically relevant field setting is relatively unknown for reptiles. Oxidative stress is a multi-faceted concept that dictates key survival and fitness parameters in any organism. Fipronil and fenitrothion are two pesticides widely used globally for agricultural pest management. Using a field-based, BACI designed experiment we investigated the impact of sublethal pesticide exposure on oxidative stress biomarkers protein carbonyl and DNA damage (8-OHdG), in an arid-zone lizard species, Pogona vitticeps. A single ecologically relevant dose of pesticide was applied via oral gavage to treatment animals. Lizard condition, activity measures, and blood biomarkers were measured at relevant sampling intervals. Cholinesterase (ChE) and acetylcholinesterase (AChE) enzymatic biomarkers were measured in response to fenitrothion, and fipronil blood residues were measured for fipronil-treated lizards. Results suggested no significant treatment effect of either pesticide on parameters measured, however, 8-OHdG levels decreased by ≥ 45% for both pesticide treatment groups and not controls. Protein carbonyl levels showed a high degree of individual variation that proved more influential than pesticide exposure. Building our understanding of the macromolecular impacts of sublethal pesticide exposure on wild lizard populations is an integral step in addressing the current gap in literature and management practices. Our study has also highlighted the complex nature of studying oxidative stress in the field and the sheer necessity of future study.

Oxidative stress and behavioral responses of moorish geckos (Tarentola mauritanica) submitted to the presence of an introduced potential predator (Hemorrhois hippocrepis).

Stressful situations induce an increase in the production of reactive oxygen species (ROS) which can lead to molecular damage and alteration of cell function. The introduction of new potential predators induces physiological stress in native fauna. However, behavioral responses have been reported in preys, demonstrating an induction of the defenses against alien species. Behavioral and antioxidant enzyme responses in the moorish gecko, Tarentola mauritanica, against the invasive predator horseshoe whip snake (Hemorrhois hippocrepis) were assessed. Behavior was recorded and a tissue sample from the tail was collected after placing the gecko in a terrarium with previous absence or presence of the snake in 'Control' and 'H. hippocrepis' groups, respectively. Fifteen behavioral variables were examined, including tongue flick (TF) and locomotion patterns. Antioxidant enzyme activities -catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), glutathione reductase (GR)-, and the levels of reduced (GSH) and oxidized glutathione (GSSG), glutathione/glutathione disulfide ratio (GSH/GSSG) and malondialdehyde (MDA) concentrations were measured in the tissue sampled. Geckos exposed to the snake's odor showed a higher number of TF, longer amounts of time remaining motionless or moving in slow motion and they spent less time on the ground in comparison to the 'Control' group. The presence of the snake produced a significant increase in the activities of CAT, SOD and GR and a decrease in the GSH/GSSG ratio in T. mauritanica individuals exposed to the snake's scent. Thus, both behavioral responses and oxidative stress biomarkers clearly showed that T. mauritanica is able to recognize H. hippocrepis as a potential predator, despite being a recently introduced snake at the Balearic Islands.

Metabolic and Hormonal Changes Associated with Vitellogenesis in Salvator merianae Lizards.

Vitellogenesis in oviparous vertebrates is a critical marker of the restart of seasonal reproductive activity. During this process of multihormonal regulation, females allocate a considerable amount of organic and mineral reserves to the synthesis of yolk, with changes in their plasma values. In this work, we determined plasma levels of various metabolites and steroid hormones throughout the reproductive cycle in females of Salvator merianae who developed vitellogenic and non-vitellogenic follicular cycles. We worked for two consecutive years with 20 adult females from the Experimental Hatchery of the Facultad de Agronomía y Zootecnia of the Universidad Nacional de Tucumán. Values of metabolites: glucose, triglycerides, cholesterol, calcium, phosphorus, albumin, total proteins, and hormones: estradiol, progesterone, and testosterone, were determined during the following stages of the annual cycle: hibernation, hibernation emergence, courtship-mating, oviposition, and incubation. Vitellogenic females showed significantly higher plasma levels of triglycerides, calcium, phosphorus, and albumin than non-vitellogenic females, mainly in the courtship-mating stage (advanced vitellogenesis). In contrast, annual cholesterol averages were lower in vitellogenic females. Glucose showed changes throughout the annual cycle regardless of the vitellogenic condition. Total proteins plasma levels had very few fluctuations during the cycle. Among the hormones studied, only testosterone showed differences related to vitellogenic condition, with higher levels in non-vitellogenic females during the entire reproductive cycle. The knowledge of these changes associated with vitellogenesis will improve zootechnical management and will allow optimizing the reproductive efficiency of Salvator lizards in captivity.

Keratin intermediate filament chains in the European common wall lizard (Podarcis muralis) and a potential keratin filament crosslinker.

On the basis of sequence homology with mammalian α-keratins, and on the criteria that the coiled-coil segments and central linker in the rod domain of these molecules must have conserved lengths if they are to assemble into viable intermediate filaments, a total of 28 Type I and Type II keratin intermediate filament chains (KIF) have been identified from the genome of the European common wall lizard (Podarcis muralis). Using the same criteria this number may be compared to 33 found here in the green anole lizard (Anole carolinensis) and 25 in the tuatara (Sphenodon punctatus). The Type I and Type II KIF genes in the wall lizard fall in clusters on chromosomes 13 and 2 respectively. Although some differences occur in the terminal domains in the KIF chains of the two lizards and tuatara, the similarities between key indicator residues - cysteine, glycine and proline - are significant. The terminal domains of the KIF chains in the wall lizard also contain sequence repeats commonly based on glycine and large apolar residues and would permit the fine tuning of physical properties when incorporated within the intermediate filaments. The H1 domain in the Type II chain is conserved across the lizards, tuatara and mammals, and has been related to its role in assembly at the 2-4 molecule level. A KIF-like chain (K80) with an extensive tail domain comprised of multiple tandem repeats has been identified as having a potential filament-crosslinking role.

Fecal adrenal hormone patterns during ovulatory and non-ovulatory reproductive cycles in female veiled chameleons (Chamaeleo calyptratus).

The relationship between the reproductive (hypothalamic-pituitary-gonadal; HPG) and adrenal (hypothalamic-pituitary-adrenal; HPA) hormone axes is complex and can vary depending on the species and environmental factors affecting an individual. In an effort to understand this relationship in female veiled chameleons (Chamaeleo calyptratus), the patterns of fecal metabolites of corticosterone (C), estradiol (E), testosterone (T), and progesterone (P) were analyzed by enzyme immunoassay (EIA) during ovulatory (OC; eggs laid) and non-ovulatory cycles (NOC; no eggs laid). Glucocorticoid (GC) metabolites in the fecal extracts were characterized by HPLC and corticosterone EIA performance was assessed by parallelism, accuracy and precision tests. The results indicated that the assay chosen reliably measured the hormone metabolites present in the fecal extracts. Regular, cyclical hormone metabolite patterns consisting of an E peak followed by peaks of T, P and C in close succession were observed during both ovulatory and non-ovulatory cycles; relative levels of P and C, however, were higher during ovulatory cycles. Corticosterone metabolite levels, in particular, increased throughout vitellogenesis and peaked in late vitellogenesis (in non-ovulatory cycles) or around the time of ovulation, and remained elevated throughout the gravid period, falling just prior to oviposition. The results provide evidence of variation in glucocorticoid production throughout different stages of the reproductive cycle, including a role in the ovulatory process; the physiology, however, remains unclear.

The nocturnal leopard gecko (Eublepharis macularius) uses UVb radiation for vitamin D3 synthesis.

Vitamin D is an important regulator of calcium and phosphorus homeostasis in animals. It can be acquired from the diet or synthesised de novo when skin is exposed to UVb. Vitamin D deficiency can lead to a complex of diseases collectively called metabolic bone disease (MBD). Diurnal lizards without access to UVb are prone to develop vitamin D deficiency, even when dietary vitamin D3 is provided. A trial was conducted to determine whether juvenile nocturnal lizards require access to UVb to prevent vitamin D deficiency. All leopard geckos (Eublepharis macularius) were supplemented with dietary vitamin D3. One group was exposed to low level UVb radiation (33-51 µW/cm2) from hatching until 6 months of age and a second group remained unexposed. Animals were fed ad libitum and their growth and weight gain compared with non-exposed controls. At the end of the trial, blood samples were analysed for vitamin D3 metabolites. The concentration of the vitamin D3 metabolite, 25(OH)D3, was higher in UVb exposed animals (61 ± 20 vs. 38 ± 8 nmol/L), confirming cutaneous synthesis with UVb exposure. Growth and weight gain were similar in both groups, and this, together with the absence of clinical symptoms, suggests that dietary vitamin D3 alone can meet the vitamin D requirements for growth of this nocturnal gecko, during the first six months of life. It remains to be investigated whether the higher vitamin D metabolite levels holds other health benefits for this species, such as improved bone density or immune response.

Venom gland transcriptome from Heloderma horridum horridum by high-throughput sequencing.

Lizards of the Helodermatidae (Anguimorpha) family consist of at least two well recognized species: Heloderma horridum horridum and Heloderma suspectum suspectum. They contain specialized glands in their jaws that produce venomous secretions that causes envenoming symptoms to bitten animals. One way to study proteins from such secretions is by RNA-seq; a powerful molecular tool to characterize the transcriptome of such specialized gland, and its protein secretions. The total RNA from venom gland tissues of H. horridum horridum was extracted and a cDNA library was constructed and sequenced. Overall, 114,172 transcripts were found, and 199 were annotated based on sequence similarities to previously described peptides/proteins. Transcripts coding for putative exendins, defensins, natriuretics and serine protease inhibitors were the most highly expressed. Transcripts that code for several putative serine proteases, phospholipases, metalloproteases, lipases, L-amino oxidase and nucleases were also found. Some of the novel identified transcripts were translationally controlled tumor proteins, venom factors, vespryns, waprins, lectins, cystatins and serine protease inhibitors. All these new protein structures may contribute to a better understanding of the venomous secretions of the Helodermatidae family.

Steroid hormones, energetic state, and immunocompetence vary across reproductive contexts in a parthenogenetic lizard.

Reproduction is energetically expensive and investing in this life history trait is likely accompanied by significant changes in physiological activity. Investment strategy necessary for achieving reproductive success in reptiles can vary with reproductive form and pattern, potentiating different consequences for competing fitness-related traits such as those key to survival. The goal of this study was to assess if and how energetic state (i.e., energy metabolites) and self-maintenance (i.e., immunocompetence) are hormonally modulated across reproductive contexts in an oviparous, parthenogenetic lizard, the Colorado Checkered Whiptail Aspidoscelis neotesselata. Here blood plasma samples were collected from lizards within the US Army Fort Carson Military Installation near Colorado Springs, CO, USA, during seasons of reproductive activity (i.e., June) and inactivity (i.e., August). Measures of reproductive (i.e., estradiol) and energy-mobilizing (i.e., corticosterone) hormones, energy metabolites (i.e., glucose, triglycerides, and free glycerol), and innate immunity (i.e., bactericidal ability) were compared by season and reproductive stage. Levels of energy metabolites and bactericidal ability were compared to levels of E2 and CORT. Bactericidal ability was also compared to levels of energy metabolites. Corticosterone and glucose levels were lower during the reproductive season while triglyceride levels and bactericidal ability were higher, but both estradiol and free glycerol levels did not differ between seasons. Throughout vitellogenesis, corticosterone and glucose levels as well as bactericidal ability did not differ, but estradiol levels were higher during early and mid-stage and both triglyceride and free glycerol levels were lower during gravidity. Corticosterone levels were negatively associated with circulating triglycerides and bactericidal ability, but were not related to glucose nor free glycerol levels. Estradiol levels were positively associated with free glycerol levels and bactericidal ability, but were not related to glucose nor triglyceride levels. Finally, bactericidal ability was negatively associated with glucose, but positively associated with triglycerides. Differences in energetic state and immunocompetence are thus reflected by shifts in hormone secretion across reproductive investment. These findings provide partial support for the hypothesis that energetic state is differentially regulated by steroid hormones to afford reproduction, potentially at the cost of future survival.

Investment of both essential fatty and amino acids to immunity varies depending on reproductive stage.

Trade-offs among the key life-history traits of reproduction and immunity have been widely documented. However, the currency in use is not well-understood. We investigated how reproducing female side-blotched lizards, Uta stansburiana, allocate lipids versus proteins when given an immune challenge. We tested whether lizards would invest more in reproduction or immunity depending on reproductive stage. Females were given stable isotopes (15 N-leucine and 13 C-1-palmitic acid), maintained on a regular diet and given either a cutaneous biopsy or a sham biopsy (control). Stable isotopes were monitored and analyzed in feces and uric acid, skin biopsies, eggs, and toe clips. We found that lizards deposited both proteins and lipids into their healing wounds (immune-challenged), skin (control), and eggs (all) and that catabolism of proteins exceeded incorporation into tissue during wound-healing. Specifically, we found that healed biopsies of wounded animals had more leucine and palmitic acid than the nonregrown skin biopsies taken from unwounded control animals. Earlier in reproduction, lizards invested relatively more labeled proteins into healing their wound tissue, but not into unwounded skin of control animals. Thus, reproduction is sometimes favored over self-maintenance, but only in later reproductive stages. Finally, we documented positive relationships among the amount of palmitic acid deposited in the eggs, the amount of food eaten, and the amount of palmitic acid excreted, suggesting higher turnover rates of lipids in lizards investing highly in their eggs.

PGE2 facilitates tail regeneration via activation of Wnt signaling in Gekko japonicus.

Tail regeneration is a distinguishing feature of lizards; however, the mechanisms underlying tail regeneration remain elusive. Prostaglandin E2 (PGE2) is an arachidonic acid metabolite that has been extensively investigated in the inflammatory response under both physiological and pathological conditions. PGE2 also act as a regulator of hematopoietic stem cell homeostasis by interacting with Wnt signaling molecules. The present study aims to identify the effects of PGE2 on tail regeneration and the molecular mechanisms behind it. We initially found that PGE2 levels increased during the early stages of tail regeneration, accompanied by the up-regulated expression of cyclooxygenase 1 and cyclooxygenase 2. Next, we demonstrated that reduced PGE2 production leads to the retardation of tail regeneration. Subsequent experiments demonstrated that this effect is likely mediated by Wnt signaling, which proposing that the activation of the Wnt pathway is essential for the initiation of regeneration. The results showed that inhibition of PGE2 production could suppress Wnt activation and inhibit the proliferation of both epithelial and blastema cells. Furthermore, our findings indicated that forced activation of Wnt signaling could rescue the inhibitory effect of Cox antagonist on regeneration, suggesting a positive role of PGE2 on tail regeneration via a non-inflammatory mechanism.

Morph-specific protein patterns in the femoral gland secretions of a colour polymorphic lizard.

Colour polymorphism occurs when two or more genetically-based colour morphs permanently coexist within an interbreeding population. Colouration is usually associated to other life-history traits (ecological, physiological, behavioural, reproductive …) of the bearer, thus being the phenotypic marker of such set of genetic features. This visual badge may be used to inform conspecifics and to drive those decision making processes which may contribute maintaining colour polymorphism under sexual selection context. The importance of such information suggests that other communication modalities should be recruited to ensure its transfer in case visual cues were insufficient. Here, for the first time, we investigated the potential role of proteins from femoral gland secretions in signalling colour morph in a polymorphic lizard. As proteins are thought to convey identity-related information, they represent the ideal cues to build up the chemical modality used to badge colour morphs. We found strong evidence for the occurrence of morph-specific protein profiles in the three main colour-morphs of the common wall lizard, which showed both qualitative and quantitative differences in protein expression. As lizards are able to detect proteins by tongue-flicking and vomeronasal organ, this result support the hypothesis that colour polymorphic lizards may use a multimodal signal to inform about colour-morph.

Gekkonidae, Lizard tail extracts elicit apoptotic response against non-small cell lung cancer via inhibiting Akt signaling.

The genes of Gekkonidae, a lizard, are known to be very similar to human genes. According to previous research, lizard extracts inhibit angiogenesis and show anticancer activity against various cancers. In this regard, this study assessed whether lizard tail extracts (LTE) cause anticancer activity against lung cancer in mouse and human lung cancer cell lines. The results showed that LTE exhibited anticancer activity against lung cancer in vitro and in vivo. In vitro, cell viability and proliferation decreased in two lung cancer cell lines, while annexin V and single-stranded DNA both increased, showing apoptotic activity caused by LTE. We also found that LTE induced apoptosis in a caspase-3/7 cascade-dependent manner and inhibited the phosphorylation of Akt by participating in the PI3k/Akt pathway. In vivo, LTE decreased tumor volume in LLC bearing mice. Our results demonstrate the potential of LTE as a natural-derived anticancer drug to overcome the chemotherapy side effects during cancer treatment and contribute to the discovery of candidate substances.

Health status of the lizard Podarcis siculus (Rafinesque-Schmaltz, 1810) subject to different anthropogenic pressures.

The terrestrial lizard Podarcis siculus is the most abundant reptile in Italy, where is considered a ubiquitous species. This lizard is widely distributed from the islands to the Apennines, from cultivated fields to anthropized areas such as gardens and city parks. For this reason, these animals are exposed to extensive physical and chemical stresses, as well as to the possibility of coming into contact with industrial pollutants and substances used in agricultural practices. Here, we review the health status of lizard specimens inhabiting natural non-anthropized areas and fields devoted to organic farming, considering the condition of (1) liver, representing the main detoxifying organ, directly influenced by feeding, and (2) gonads, essential for reproduction and, therefore, for the survival of the species. The morphological and biomolecular condition of these organs was then compared with those obtained from lizard specimens experimentally treated with nonylphenol, a co-formulant of many insecticides and plant protection products widely used in conventional farming, known to have harmful estrogenic effects. Taken together, data demonstrate that lizards inhabiting manured soil are in good health status and show a regular morphology of liver, testis, and ovary. Animals are found to be less exposed to the toxic heavy metals cadmium and lead if compared with specimens collected in areas not devoted to agriculture, but probably more exposed to vehicular traffic. However, manure, as well as nonylphenol, exerts a xeno-estrogenic effect, particularly evident in male specimens, more sensitive to estrogenic contamination.

Immunohistochemical staining patterns of alpha-keratins in normal tissues from two reptile species: implications for characterization of squamous cell carcinomas.

BACKGROUND: Cytokeratins with epitopes in common with those of alpha (acidic and basic) mammalian keratins have been immunohistochemically demonstrated in the epidermis of reptiles. However, there are no reports of immunohistochemical staining patterns of alpha-keratins in other tissues from reptiles. Because the epithelial tumours usually retain the keratin patterns of their normal epithelial origin, it is necessary to know in advance these patterns in the major normal epithelia and organs. We used anti-alpha human keratin AE1 and AE3 monoclonal antibodies to study the staining patterns of alpha-keratins in the major normal epithelia and organs from two reptile species [the bearded dragon (Pogona vitticeps) and the loggerhead sea turtle (Caretta caretta)]. We also studied the immunolocalization of alpha-keratins in squamous cell carcinomas (SCCs) in a bearded dragon and two loggerhead turtles. RESULTS: Acidic alpha-keratin (AE1 positive) was detected in many of the epithelial tissues of the bearded dragons; however, the detection of basic alpha-keratin (AE3 positive) was much more limited. Alpha-keratins were detected in a greater number of tissues of loggerhead turtles compared with those observed in bearded dragons. In the bearded dragon SCC, all layers of the nests of neoplastic cells, including the cornified layer of the keratin pearls, were strongly reactive with the AE1 antibody. However, a weak reactivity using the AE3 antibody was detected in the basal and intermediate layers of these nests. In the cutaneous SCCs of both sea turtles, acidic alpha-keratin was detected in the basal and suprabasal layers, and in all of the invasive neoplastic cords, while basic alpha-keratin was mainly detected in the invasive neoplastic cords. The pattern observed in the metastases in both turtles consisted of immunohistological detection of acidic alpha-keratin in all metastatic foci, and limited or lack of detection of basic alpha-keratin. CONCLUSIONS: This study provides, for the first time, information about the immunohistochemical staining patterns of alpha-keratins in normal tissues from bearded dragons and loggerhead sea turtles, and confirms the usefulness of AE1 and AE3 monoclonal antibodies in these reptile species. The use of these antibodies also contributed to a better characterization of SCCs in these species.