Teratogenic Toxicity Evaluation of Bladder Cancer-Specific Oncolytic Adenovirus on Mice.

BACKGROUND: In our previous studies, we had demonstrated the efficiency and specificity of constructed bladder tissue-specific adenovirus Ad-PSCAE-UPII-E1A-AR (APU-EIA-AR) on bladder cancer. The virus biodistribution and body toxicity in nude mice have also been investigated. However, the safety of the bladder cancer-specific oncolytic adenovirus on fetal mice and F1 mice should be under intense investigation. OBJECTIVE: In order to evaluate the teratogenic toxicity of bladder cancer-specific oncolytic adenovirus APU-EIA-AR on mice, in this study, we investigated the fetal mice weight, fetal body length and tail length, fetal skeleton development, as well as the F1 mice weight, growth curve, and major organ pathology. These teratogenic toxicity data of bladder tissue-specific adenovirus Ad-PSCAE- UPII-E1A-AR (AD) would provide safe information prior to embarking on clinical trials. METHODS: On the sixth day of being fertilized, the pregnant mice began to be intramuscularly administrated with AD (1×107VP, 1×108VP, 1×109VP) every other day for ten days. The pregnant mice were then divided into two groups. One group was euthanized on the seventeenth day; the fetal mice were taken out, and the bone structure of the infants was observed. The other group was observed until natural childbirth. The Filial Generation (F1) is fed for 30 days; the variations in the growth progress and development were assessed. The mice were then euthanized; The tissues from major organs were harvested and observed under the microscope. RESULTS: In the process of teratogenic toxicity test, the Placenta weight, fetal mice weight, body length, and a tail length of mice fetal in adenovirus treated group did not reveal any alteration. Meanwhile, comparing with the PBS group, there is no obvious change in the skeleton of fetal mice treated with adenovirus. During the development process of F1 mice treated with adenovirus, the changes in mice weight show statistical significance. However, in the progress of the growth curve, this difference is not very obvious. Furthermore, the pathological section showed no obvious alteration in major organs. CONCLUSION: Our study demonstrated that bladder cancer-specific adenovirus Ad-PSCAE-UPII- E1A-AR appears safe in pregnant mice without any discernable effects on fetal mice and F1 development. Hence, it is relatively safe for tumor gene therapy.

Elimination of Teratogenic Human Induced Pluripotent Stem Cells by Bee Venom via Calcium-Calpain Pathway.

Induced pluripotent stem cells (iPSCs) are regarded as a promising option for cell-based regenerative medicine. To obtain safe and efficient iPSC-based cell products, it is necessary to selectively eliminate the residual iPSCs prior to in vivo implantation due to the risk of teratoma formation. Bee venom (BV) has long been used in traditional Chinese medicine to treat inflammatory diseases and relieve pain, and has been shown to exhibit anti-cancer, anti-mutagenic, anti-nociceptive, and radioprotective activities. However, the potential benefits of BV in iPSC therapy, particularly its anti-teratoma activity, have not been examined. In this study, we found that BV selectively induced cell death in iPSCs, but not in iPSC-derived differentiated cells (iPSCs-Diff). BV rapidly disrupted cell membrane integrity and focal adhesions, followed by induction of apoptosis and necroptosis in iPSCs. We also found that BV remarkably enhanced intracellular calcium levels, calpain activation, and reactive oxygen speciesgeneration in iPSCs. BV treatment before in ovo grafting efficiently prevented iPSC-derived teratoma formation. In contrast, no DNA damage was observed in iPSCs-Diff following BV treatment, further demonstrating the safety of BV for use with iPSCs-Diff. Taken together, these findings show that BV has potent anti-teratoma activity by eliminating residual iPSCs, and can be used for the development of effective and safe iPSC-based cell therapies.

Valproic acid increases NF-κB transcriptional activation despite decreasing DNA binding ability in P19 cells, which may play a role in VPA-initiated teratogenesis.

The nuclear factor-kappa B (NF-κB) family of transcription factors regulate gene expression in response to diverse stimuli. We previously demonstrated that valproic acid (VPA) exposure in utero decreases total cellular protein expression of the NF-κB subunit p65 in CD-1 mouse embryos with a neural tube defect but not in phenotypically normal littermates. This study evaluated p65 mRNA and protein expression in P19 cells and determined the impact on DNA binding ability and activity. Exposure to 5mM VPA decreased p65 mRNA and total cellular protein expression however, nuclear p65 protein expression was unchanged. VPA reduced NF-κB DNA binding and nuclear protein of the p65 DNA-binding partner, p50. NF-κB transcriptional activity was increased with VPA alone, despite decreased phosphorylation of p65 at Ser276, and when combined with tissue necrosis factor α. These results demonstrate that VPA increases NF-κB transcriptional activity despite decreasing DNA binding, which may play a role in VPA-initiated teratogenesis.

Arrested Development: Infantile Hemangioma and the Stem Cell Teratogenic Hypothesis.

BACKGROUND: Early-life programming is defined by the adaptive changes made by the fetus in response to an adverse in utero environment. Infantile hemangioma (IH), a vascular anomaly, is the most common tumor of infancy. Here we take IH as the tumor model to propose the stem cell teratogenic hypothesis of tumorigenesis and the potential involvement of the immune system. OBJECTIVES: Teratogenic agents include chemicals, heavy metals, pathogens, and ionizing radiation. To investigate the etiology and pathogenesis of IH, we hypothesized that they result from a teratogenic mechanism. Immature, incompletely differentiated, dysregulated progenitor cells (multipotential stem cells) are arrested in development with vasculogenic, angiogenic, and tumorigenic potential due to exposure to teratogenic agents such as extrinsic factors that disrupt intrinsic factors via molecular mimicry. During the critical period of immunological tolerance, environmental exposure to immunotoxic agents may harness the teratogenic potential in the developing embryo or fetus and modify the early-life programming algorithm by altering normal fetal development, causing malformations, and inducing tumorigenesis. Specifically, exposure to environmental agents may interfere with physiological signaling pathways and contribute to the generation of IH, by several mechanisms. DISCUSSION: An adverse in utero environment no longer serves as a sustainable environment for proper embryogenesis and normal development. Targeted disruption of stem cells by extrinsic factors can alter the genetic program. CONCLUSIONS: This article offers new perspectives to stimulate discussion, explore novel experimental approaches (such as immunotoxicity/vasculotoxicity assays and novel isogenic models), and to address the questions raised to convert the hypotheses into nontoxic, noninvasive treatments.

Peroxisome Proliferator-Activated Receptor α Activation Is Not the Main Contributor to Teratogenesis Elicited by Polar Compounds from Oxidized Frying Oil.

We previously reported that polar compounds (PO) in cooking oil are teratogenic and perturbed retinoic acid (RA) metabolism. Considering PO as a potent peroxisome proliferator-activated receptor α (PPARα) activator, this study aimed to investigate the role of PPARα in PO-induced teratogenesis and disturbance of RA metabolism. Female PPARα knockout or wild type mice were mated with males of the same genotype. Pregnant mice were fed a diet containing 10% fat from either fresh oil (FO) or PO from gestational day1 to day18, and killed at day18. The PO diet significantly increased the incidence of teratogenesis and fetal RA concentrations, regardless of genotype. Though PPARα deficiency disturbed maternal RA homeostasis, itself did not contribute to teratogenesis as long as FO diet was given. The mRNA profile of genes involved in RA metabolism was differentially affected by diet or genotype in mothers and fetuses. Based on hepatic mRNA levels of genes involved in xenobiotic metabolism, we inferred that PO not only activated PPARα, but also altered transactivity of other xenobiotic receptors. We concluded that PO-induced fetal anomalies and RA accumulation were independent of PPARα activation.

Angiogenesis-related genes and thalidomide teratogenesis in humans: an approach on genetic variation and review of past in vitro studies.

Thalidomide embryopathy (TE) has affected more than 10,000 babies worldwide. The hypothesis of antiangiogenesis as the teratogenic mechanism of thalidomide has been investigated in several experimental models; but, in humans, it has only been accessed by in vitro studies. Here, we hypothesized the effect of thalidomide upon angiogenesis-related molecules or proteins, previously identified in human embryonic cells, through the in silico STRING-tool. We also investigated ten polymorphisms in angiogenesis-related genes in 38 Brazilian TE individuals and 136 non-affected Brazilians. NOS2, PTGS2, and VEGFA polymorphisms were chosen for genotyping. The STRING-tool suggested nitric oxide and ß-catenin as the central angiogenesis-related molecules affected by thalidomide's antiangiogenic property. We did not identify a significant difference of allelic, genotypic or haplotypic frequencies between the groups. We could not predict a risk allele or a protective one for TE in NOS2, PTGS2, or VEGFA, although other genes should be analyzed in larger samples. The role of nitric oxide and ß-catenin must be further evaluated, regarding thalidomide teratogenesis complex etiology.

Immunomodulatory drugs disrupt the cereblon-CD147-MCT1 axis to exert antitumor activity and teratogenicity.

Immunomodulatory drugs (IMiDs), such as thalidomide and its derivatives lenalidomide and pomalidomide, are key treatment modalities for hematologic malignancies, particularly multiple myeloma (MM) and del(5q) myelodysplastic syndrome (MDS). Cereblon (CRBN), a substrate receptor of the CRL4 ubiquitin ligase complex, is the primary target by which IMiDs mediate anticancer and teratogenic effects. Here we identify a ubiquitin-independent physiological chaperone-like function of CRBN that promotes maturation of the basigin (BSG; also known as CD147) and solute carrier family 16 member 1 (SLC16A1; also known as MCT1) proteins. This process allows for the formation and activation of the CD147-MCT1 transmembrane complex, which promotes various biological functions, including angiogenesis, proliferation, invasion and lactate export. We found that IMiDs outcompete CRBN for binding to CD147 and MCT1, leading to destabilization of the CD147-MCT1 complex. Accordingly, IMiD-sensitive MM cells lose CD147 and MCT1 expression after being exposed to IMiDs, whereas IMiD-resistant cells retain their expression. Furthermore, del(5q) MDS cells have elevated CD147 expression, which is attenuated after IMiD treatment. Finally, we show that BSG (CD147) knockdown phenocopies the teratogenic effects of thalidomide exposure in zebrafish. These findings provide a common mechanistic framework to explain both the teratogenic and pleiotropic antitumor effects of IMiDs.

ASK1 mediates the teratogenicity of diabetes in the developing heart by inducing ER stress and inhibiting critical factors essential for cardiac development.

Maternal diabetes in mice induces heart defects similar to those observed in human diabetic pregnancies. Diabetes enhances apoptosis and suppresses cell proliferation in the developing heart, yet the underlying mechanism remains elusive. Apoptosis signal-regulating kinase 1 (ASK1) activates the proapoptotic c-Jun NH2-terminal kinase 1/2 (JNK1/2) leading to apoptosis, suggesting a possible role of ASK1 in diabetes-induced heart defects. We aimed to investigate whether ASK1 is activated in the heart and whether deleting the Ask1 gene blocks diabetes-induced adverse events and heart defect formation. The ASK1-JNK1/2 pathway was activated by diabetes. Deleting Ask1 gene significantly reduced the rate of heart defects, including ventricular septal defects (VSDs) and persistent truncus arteriosus (PTA). Additionally, Ask1 deletion diminished diabetes-induced JNK1/2 phosphorylation and its downstream transcription factors and endoplasmic reticulum (ER) stress markers. Consistent with this, caspase activation and apoptosis were blunted. Ask1 deletion blocked the increase in cell cycle inhibitors (p21 and p27) and the decrease in cyclin D1 and D3 and reversed diabetes-repressed cell proliferation. Ask1 deletion also restored the expression of BMP4, NKX2.5, and GATA5, Smad1/5/8 phosphorylation, whose mutations or deletion result in reduced cell proliferation, VSD, and PTA formation. We conclude that ASK1 may mediate the teratogenicity of diabetes through activating the JNK1/2-ER stress pathway and inhibiting cell cycle progression, thereby impeding the cardiogenesis pathways essential for ventricular septation and outflow tract development.