Tobacco Smoke Condensate Induces Morphologic Changes in Human Papillomavirus-Positive Cervical Epithelial Cells Consistent with Epithelial to Mesenchymal Transition (EMT) with Activation of Receptor Tyrosine Kinases and Regulation of TGFB.

High-risk human papillomavirus (HR-HPV; HPV-16) and cigarette smoking are associated with cervical cancer (CC); however, the underlying mechanism(s) remain unclear. Additionally, the carcinogenic components of tobacco have been found in the cervical mucus of women smokers. Here, we determined the effects of cigarette smoke condensate (CSC; 3R4F) on human ectocervical cells (HPV-16 Ect/E6E7) exposed to CSC at various concentrations (10-6-100 µg/mL). We found CSC (10-3 or 10 µg/mL)-induced proliferation, enhanced migration, and histologic and electron microscopic changes consistent with EMT in ectocervical cells with a significant reduction in E-cadherin and an increase in the vimentin expression compared to controls at 72 h. There was increased phosphorylation of receptor tyrosine kinases (RTKs), including Eph receptors, FGFR, PDGFRA/B, and DDR2, with downstream Ras/MAPK/ERK1/2 activation and upregulation of common EMT-related genes, TGFB SNAI2, PDGFRB, and SMAD2. Our study demonstrated that CSC induces EMT in ectocervical cells with the upregulation of EMT-related genes, expression of protein biomarkers, and activation of RTKs that regulate TGFB expression, and other EMT-related genes. Understanding the molecular pathways and environmental factors that initiate EMT in ectocervical cells will help delineate molecular targets for intervention and define the role of EMT in the initiation and progression of cervical intraepithelial neoplasia and CC.

Differential roles of serotonin receptor subtypes in regulation of neurotrophin receptor expression and intestinal hypernociception.

OBJECTIVES: Aberrant serotonin (5-hydroxytryptamine, 5-HT) metabolism and neurite outgrowth were associated with abdominal pain in irritable bowel syndrome (IBS). We previously demonstrated that 5-HT receptor subtype 7 (5-HT7) was involved in visceral hypersensitivity of IBS-like mouse models. The aim was to compare the analgesic effects of a novel 5-HT7 antagonist to reference standards in mouse models and investigate the mechanisms of 5-HT7-dependent neuroplasticity. METHODS: Two mouse models, including Giardia post-infection combined with water avoidance stress (GW) and post-resolution of trinitrobenzene sulfonic acid-induced colitis (PT) were used. Mice were orally administered CYY1005 (CYY, a novel 5-HT7 antagonist), alosetron (ALN, a 5-HT3 antagonist), and loperamide (LPM, an opioid receptor agonist) prior to measurement of visceromotor responses (VMR). Levels of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin receptors (NTRs) were assessed. RESULTS: Peroral CYY was more potent than ALN or LPM in reducing VMR values in GW and PT mice. Increased mucosal 5-HT7-expressing nerve fibers were associated with elevated Gap43 levels in the mouse colon. We observed higher colonic Ntrk2 and Ngfr expression in GW mice, and increased Bdnf expression in PT mice compared with control mice. Human SH-SY5Y cells stimulated with mouse colonic supernatant or exogenous serotonin exhibited longer nerve fibers, which CYY dose-dependently inhibited. Serotonin increased Ntrk1 and Ngfr expression via 5-HT7 but not 5-HT3 or 5-HT4, while Ntrk2 upregulation was dependent on all three 5-HT receptor subtypes. CONCLUSIONS: Stronger analgesic effects by peroral CYY were observed compared with reference standards in two IBS-like mouse models. The 5-HT7-dependent NTR upregulation and neurite elongation may be involved in intestinal hypernociception.

Design, Synthesis, and Evaluation of Glucose Transporter Inhibitor-SN38 Conjugates for Targeting Colorectal Cancer.

Irinotecan (1), a prodrug of SN38 (2) approved by the US Food and Drug Administration for treating colorectal cancer, lacks specificity and causes many side effects. To increase the selectivity and therapeutic efficacy of this drug, we designed and synthesized conjugates of SN38 and glucose transporter inhibitors (phlorizin (5) or phloretin (6)), which could be hydrolyzed by glutathione or cathepsin to release SN38 in the tumor microenvironment, as a proof of concept. These conjugates (8, 9, and 10) displayed better antitumor efficacy with lower systemic exposure to SN38 in an orthotopic colorectal cancer mouse model compared with irinotecan at the same dosage. Further, no major adverse effects of the conjugates were observed during treatment. Biodistribution studies showed that conjugate 10 could induce higher concentrations of free SN38 in tumor tissues than irinotecan at the same dosage. Thus, the developed conjugates exhibit potential for treating colorectal cancer.

Transepithelial Barrier Dysfunction Drives Microbiota Dysbiosis to Initiate Epithelial Clock-driven Inflammation.

BACKGROUND: Factors that contribute to inflammatory bowel disease [IBD] pathogenesis include genetic polymorphisms, barrier loss, and microbial dysbiosis. A major knowledge gap exists in the origins of the colitogenic microbiome and its relationship with barrier impairment. Epithelial myosin light chain kinase [MLCK] is a critical regulator of the paracellular barrier, but the effects of MLCK activation on the intraepithelial bacteria [IEB] and dysbiosis are incompletely understood. We hypothesise that MLCK-dependent bacterial endocytosis promotes pathobiont conversion and shapes a colitogenic microbiome. METHODS: To explore this, transgenic [Tg] mice with barrier loss induced by intestinal epithelium-specific expression of a constitutively active MLCK were compared with wild-type [WT] mice. RESULTS: When progeny of homozygous MLCK-Tg mice were separated after weaning by genotype [Tg/Tg, Tg/WT, WT/WT], increased IEB numbers associated with dysbiosis and more severe colitis were present in Tg/Tg and Tg/WT mice, relative to WT/WT mice. Cohousing with MLCK-Tg mice induced dysbiosis, increased IEB abundance, and exacerbated colitis in WT mice. Conversely, MLCK-Tg mice colonised with WT microbiota at birth displayed increased Escherichia abundance and greater colitis severity by 6 weeks of age. Microarray analysis revealed circadian rhythm disruption in WT mice co-housed with MLCK-Tg mice relative to WT mice housed only with WT mice. This circadian disruption required Rac1/STAT3-dependent microbial invasion but not MLCK activity, and resulted in increased proinflammatory cytokines and glucocorticoid downregulation. CONCLUSIONS: The data demonstrate that barrier dysfunction induces dysbiosis and expansion of invasive microbes that lead to circadian disruption and mucosal inflammation. These results suggest that barrier-protective or bacterium-targeted precision medicine approaches may be of benefit to IBD patients.

Photoinduced electron transfer in cytochrome bc1: Dynamics of rotation of the Iron-sulfur protein during bifurcated electron transfer from ubiquinol to cytochrome c1 and cytochrome bL.

The electron transfer reactions within wild-type Rhodobacter sphaeroides cytochrome bc1 (cyt bc1) were studied using a binuclear ruthenium complex to rapidly photooxidize cyt c1. When cyt c1, the iron­sulfur center Fe2S2, and cyt bH were reduced before the reaction, photooxidation of cyt c1 led to electron transfer from Fe2S2 to cyt c1 with a rate constant of ka = 80,000 s-1, followed by bifurcated reduction of both Fe2S2 and cyt bL by QH2 in the Qo site with a rate constant of k2 = 3000 s-1. The resulting Q then traveled from the Qo site to the Qi site and oxidized one equivalent each of cyt bL and cyt bH with a rate constant of k3 = 340 s-1. The rate constant ka was decreased in a nonlinear fashion by a factor of 53 as the viscosity was increased to 13.7. A mechanism that is consistent with the effect of viscosity involves rotational diffusion of the iron­sulfur protein from the b state with reduced Fe2S2 close to cyt bL to one or more intermediate states, followed by rotation to the final c1 state with Fe2S2 close to cyt c1, and rapid electron transfer to cyt c1.

Individuals with ventromedial frontal damage display unstable but transitive preferences during decision making.

The ventromedial frontal lobe (VMF) is important for decision-making, but the precise causal role of the VMF in the decision process has not been fully established. Previous studies have suggested that individuals with VMF damage violate transitivity, a hallmark axiom of rational decisions. However, these prior studies cannot properly distinguish whether individuals with VMF damage are truly prone to choosing irrationally from whether their preferences are simply more variable. We had individuals with focal VMF damage, individuals with other frontal damage, and healthy controls make repeated choices across three categories-artworks, chocolate bar brands, and gambles. Using proper tests of transitivity, we find that, in our study, individuals with VMF damage make rational decisions consistent with transitive preferences, even though they exhibit greater variability in their preferences. That is, the VMF is necessary for having strong and reliable preferences, but not for being a rational decision maker. VMF damage affects the variability with which value is assessed, but not the consistency with which value is sought.

Glucose Conferred Irinotecan Chemoresistance through Divergent Actions of Pyruvate and ATP in Cell Death and Proliferation of Colorectal Cancer.

BACKGROUND: Altered glucose metabolism is associated with chemoresistance in colorectal cancer (CRC). This study aimed to illustrate the molecular mechanisms of glucose-mediated chemoresistance against irinotecan, a topoisomerase I inhibitor, focusing on the distinct roles of metabolites such as pyruvate and ATP in modulating cell death and proliferation. METHODS: Four human CRC cell lines, tumorspheres, and mouse xenograft models were treated with various doses of irinotecan in the presence of various concentrations of glucose, pyruvate, or ATP-encapsulated liposomes. RESULTS: In this study, human CRC cell lines treated with irinotecan in high glucose displayed increased cell viability and larger xenograft tumor sizes in mouse models compared to those treated in normal glucose concentrations. Irinotecan induced apoptosis and necroptosis, both mitigated by high glucose. Liposomal ATP prevented irinotecan-induced apoptosis, while it did not affect necroptosis. In contrast, pyruvate attenuated the receptor-interacting protein kinase 1/3-dependent necroptosis via free radical scavenging without modulating apoptotic levels. Regarding the cell cycle, liposomal ATP aggravated the irinotecan-induced G0/G1 shift, whereas pyruvate diminished the G0/G1 shift, showing opposite effects on proliferation. Last, tumorsphere structural damage, an index of solid tumor responsiveness to chemotherapy, was determined. Liposomal ATP increased tumorsphere size while pyruvate prevented the deformation of spheroid mass. CONCLUSIONS: Glucose metabolites confer tumor chemoresistance via multiple modes of action. Glycolytic pyruvate attenuated irinotecan-induced necroptosis and potentiated drug insensitivity by shifting cells from a proliferative to a quiescent state. On the other hand, ATP decreased irinotecan-induced apoptosis and promoted active cell proliferation, contributing to tumor recurrence. Our findings challenged the traditional view of ATP as the main factor for irinotecan chemoresistance and provided novel insights of pyruvate acting as an antioxidant responsible for drug insensitivity, which may shed light on the development of new therapies against recalcitrant cancers.

5-HT7 receptor-dependent intestinal neurite outgrowth contributes to visceral hypersensitivity in irritable bowel syndrome.

Irritable bowel syndrome (IBS) is characterized by visceral hypersensitivity (VH) associated with abnormal serotonin/5-hydroxytryptamine (5-HT) metabolism and neurotrophin-dependent mucosal neurite outgrowth. The underlying mechanisms of VH remain poorly understood. We investigated the role of 5-HT7 receptor in mucosal innervation and intestinal hyperalgesia. A high density of mucosal nerve fibres stained for 5-HT7 was observed in colonoscopic biopsy specimens from IBS patients compared with those from healthy controls. Staining of 5-HT3 and 5-HT4 receptors was observed mainly in colonic epithelia with comparable levels between IBS and controls. Visceromotor responses to colorectal distension were evaluated in two mouse models, one postinfectious with Giardia and subjected to water avoidance stress (GW) and the other postinflammatory with trinitrobenzene sulfonic acid-induced colitis (PT). Increased VH was associated with higher mucosal density of 5-HT7-expressing nerve fibres and elevated neurotrophin and neurotrophin receptor levels in the GW and PT mice. The increased VH was inhibited by intraperitoneal injection of SB-269970 (a selective 5-HT7 antagonist). Peroral multiple doses of CYY1005 (a novel 5-HT7 ligand) decreased VH and reduced mucosal density of 5-HT7-expressing nerve fibres in mouse colon. Human neuroblastoma SH-SY5Y cells incubated with bacteria-free mouse colonic supernatant, 5-HT, nerve growth factor, or brain-derived neurotrophic factor exhibited nerve fibre elongation, which was inhibited by 5-HT7 antagonists. Gene silencing of HTR7 also reduced the nerve fibre length. Activation of 5-HT7 upregulated NGF and BDNF gene expression, while stimulation with neurotrophins increased the levels of tryptophan hydroxylase 2 and 5-HT7 in neurons. A positive-feedback loop was observed between serotonin and neurotrophin pathways via 5-HT7 activation to aggravate fibre elongation, whereby 5-HT3 and 5-HT4 had no roles. In conclusion, 5-HT7-dependent mucosal neurite outgrowth contributed to VH. A novel 5-HT7 antagonist could be used as peroral analgesics for IBS-related pain.

Prolonged Cadmium Exposure Alters Migration Dynamics and Increases Heterogeneity of Human Uterine Fibroid Cells-Insights from Time Lapse Analysis.

Cadmium (Cd) is one of the most prevalent environmental heavy metal contaminants and is considered an endocrine disruptor and carcinogen. In women with uterine fibroids, there is a correlation between blood Cd levels and fibroid tumor size. In this study, fibroid cells were exposed to 10 µM CdCl2 for 6 months and a fast-growing Cd-Resistant Leiomyoma culture, termed CR-LM6, was recovered. To characterize the morphological and mechanodynamic features of uterine fibroid cells associated with prolonged Cd exposure, we conducted time lapse imaging using a Zeiss confocal microscope and analyzed data by Imaris and RStudio. Our experiments recorded more than 64,000 trackable nuclear surface objects, with each having multiple parameters such as nuclear size and shape, speed, location, orientation, track length, and track straightness. Quantitative analysis revealed that prolonged Cd exposure significantly altered cell migration behavior, such as increased track length and reduced track straightness. Cd exposure also significantly increased the heterogeneity in nuclear size. Additionally, Cd significantly increased the median and variance of instantaneous speed, indicating that Cd exposure results in higher speed and greater variation in motility. Profiling of mRNA by NanoString analysis and Ingenuity Pathway Analysis (IPA) strongly suggested that the direction of gene expression changes due to Cd exposure enhanced cell movement and invasion. The altered expression of extracellular matrix (ECM) genes such as collagens, matrix metallopeptidases (MMPs), secreted phosphoprotein 1 (SPP1), which are important for migration contact guidance, may be responsible for the greater heterogeneity. The significantly increased heterogeneity of nuclear size, speed, and altered migration patterns may be a prerequisite for fibroid cells to attain characteristics favorable for cancer progression, invasion, and metastasis.

Assessing the Health Outcomes of the Food Access Pilot Project: An Evaluation of a Medically Supportive Food Support Program for People Living with HIV in Rural California Counties.

Food insecurity disproportionately affects rural communities and people living with HIV (PLHIV). The Food Access Pilot Project (FAPP) was a California state-funded program that provided home-delivered, medically supportive meals via online meal vendors to food-insecure PLHIV in three rural counties. We performed longitudinal, retrospective analyses of FAPP participant data (n = 158; 504 and 460 person-time observations for viral load and CD4 count, respectively) over 36 months from a Ryan White client management database. Pre-post analyses demonstrated increased prevalence of food security and CD4 ≥ 500 between baseline and 12 months. Population-averaged trends using generalized estimating equations adjusted for participant demographics demonstrated increased odds of viral suppression and CD4 ≥ 500, and increased CD4 count (cells/mm3) for every six months of program enrollment. Home-delivered, medically supportive meals may improve food security status, HIV viral suppression, and immune health for low-income PLHIV in rural settings.

Short-term tetrabromobisphenol A exposure promotes fibrosis of human uterine fibroid cells in a 3D culture system through TGF-beta signaling.

Tetrabromobisphenol A (TBBPA), a derivative of BPA, is a ubiquitous environmental contaminant with weak estrogenic properties. In women, uterine fibroids are highly prevalent estrogen-responsive tumors often with excessive accumulation of extracellular matrix (ECM) and may be the target of environmental estrogens. We have found that BPA has profibrotic effects in vitro, in addition to previous reports of the in vivo fibrotic effects of BPA in mouse uterus. However, the role of TBBPA in fibrosis is unclear. To investigate the effects of TBBPA on uterine fibrosis, we developed a 3D human uterine leiomyoma (ht-UtLM) spheroid culture model. Cell proliferation was evaluated in 3D ht-UtLM spheroids following TBBPA (10-6 -200 µM) administration at 48 h. Fibrosis was assessed using a Masson's Trichrome stain and light microscopy at 7 days of TBBPA (10-3  µM) treatment. Differential expression of ECM and fibrosis genes were determined using RT² Profiler™ PCR arrays. Network and pathway analyses were conducted using Ingenuity Pathway Analysis. The activation of pathway proteins was analyzed by a transforming growth factor-beta (TGFB) protein array. We found that TBBPA increased cell proliferation and promoted fibrosis in 3D ht-UtLM spheroids with increased deposition of collagens. TBBPA upregulated the expression of profibrotic genes and corresponding proteins associated with the TGFB pathway. TBBPA activated TGFB signaling through phosphorylation of TGFBR1 and downstream effectors-small mothers against decapentaplegic -2 and -3 proteins (SMAD2 and SMAD3). The 3D ht-UtLM spheroid model is an effective system for studying environmental agents on human uterine fibrosis. TBBPA can promote fibrosis in uterine fibroid through TGFB/SMAD signaling.

Invasive Pathobionts Contribute to Colon Cancer Initiation by Counterbalancing Epithelial Antimicrobial Responses.

BACKGROUND & AIMS: Microbiota dysbiosis and mucosa-associated bacteria are involved in colorectal cancer progression. We hypothesize that an interaction between virulent pathobionts and epithelial defense promotes tumorigenesis. METHODS: Chemical-induced CRC mouse model was treated with antibiotics at various phases. Colonic tissues and fecal samples were collected in a time-serial mode and analyzed by gene microarray and 16S rRNA sequencing. Intraepithelial bacteria were isolated using a gentamicin resistance assay, and challenged in epithelial cultures. RESULTS: Our study showed that antibiotic treatment at midphase but not early or late phase reduced mouse tumor burden, suggesting a time-specific host-microbe interplay. A unique antimicrobial transcriptome profile showing an inverse relationship between autophagy and oxidative stress genes was correlated with a transient surge in microbial diversity and virulence emergence in mouse stool during cancer initiation. Gavage with fimA/fimH/htrA-expressing invasive Escherichia coli isolated from colonocytes increased tumor burden in recipient mice, whereas inoculation of bacteria deleted of htrA or triple genes did not. The invasive E.coli suppressed epithelial autophagy activity through reduction of microtubule-associated protein 1 light-chain 3 transcripts and caused dual oxidase 2-dependent free radical overproduction and tumor cell hyperproliferation. A novel alternating spheroid culture model was developed for sequential bacterial challenge to address the long-term changes in host-microbe interaction for chronic tumor growth. Epithelial cells with single bacterial encounter showed a reduction in transcript levels of autophagy genes while those sequentially challenged with invasive E.coli showed heightened autophagy gene expression to eliminate intracellular microbes, implicating that bacteria-dependent cell hyperproliferation could be terminated at late phases. Finally, the presence of bacterial htrA and altered antimicrobial gene expression were observed in human colorectal cancer specimens. CONCLUSIONS: Invasive pathobionts contribute to cancer initiation during a key time frame by counterbalancing autophagy and oxidative stress in the colonic epithelium. Monitoring gut microbiota and antimicrobial patterns may help identify the window of opportunity for intervention with bacterium-targeted precision medicine.

Genistein: Dual Role in Women's Health.

Advanced research in recent years has revealed the important role of nutrients in the protection of women's health and in the prevention of women's diseases. Genistein is a phytoestrogen that belongs to a class of compounds known as isoflavones, which structurally resemble endogenous estrogen. Genistein is most often consumed by humans via soybeans or soya products and is, as an auxiliary medicinal, used to treat women's diseases. In this review, we focused on analyzing the geographic distribution of soybean and soya product consumption, global serum concentrations of genistein, and its metabolism and bioactivity. We also explored genistein's dual effects in women's health through gathering, evaluating, and summarizing evidence from current in vivo and in vitro studies, clinical observations, and epidemiological surveys. The dose-dependent effects of genistein, especially when considering its metabolites and factors that vary by individuals, indicate that consumption of genistein may contribute to beneficial effects in women's health and disease prevention and treatment. However, consumption and exposure levels are nuanced because adverse effects have been observed at lower concentrations in in vitro models. Therefore, this points to the duplicity of genistein as a possible therapeutic agent in some instances and as an endocrine disruptor in others.

Adaptive learning is structure learning in time.

People use information flexibly. They often combine multiple sources of relevant information over time in order to inform decisions with little or no interference from intervening irrelevant sources. They adjust the degree to which they use new information over time rationally in accordance with environmental statistics and their own uncertainty. They can even use information gained in one situation to solve a problem in a very different one. Learning flexibly rests on the ability to infer the context at a given time, and therefore knowing which pieces of information to combine and which to separate. We review the psychological and neural mechanisms behind adaptive learning and structure learning to outline how people pool together relevant information, demarcate contexts, prevent interference between information collected in different contexts, and transfer information from one context to another. By examining all of these processes through the lens of optimal inference we bridge concepts from multiple fields to provide a unified multi-system view of how the brain exploits structure in time to optimize learning.

Subjective value, not a gridlike code, describes neural activity in ventromedial prefrontal cortex during value-based decision-making.

Across many studies, ventromedial prefrontal cortex (vmPFC) activity has been found to correlate with subjective value during value-based decision-making. Recently, however, vmPFC has also been shown to reflect a hexagonal gridlike code during navigation through physical and conceptual space, and such gridlike codes have been proposed to enable value-based choices between novel options. Here, we first show that, in theory, a hexagonal gridlike code can in some cases mimic vmPFC activity previously attributed to subjective value, raising the possibility that the subjective value correlates previously observed in vmPFC may have actually been a misconstrued gridlike signal. We then compare the two accounts empirically, using fMRI data from a large number of subjects performing an intertemporal choice task. We find clear and unambiguous evidence that subjective value is a better description of vmPFC activity in this task than a hexagonal gridlike code. In fact, we find no significant evidence at all for a hexagonal gridlike code in vmPFC activity during intertemporal choice. This result limits the generality of gridlike modulation as description of vmPFC activity. We suggest that vmPFC may flexibly switch representational schemes so as to encode the most relevant information for the current task.

A novel tumor suppressor role of myosin light chain kinase splice variants through downregulation of the TEAD4/CD44 axis.

Myosin light chain kinase (MLCK) regulates actinomyosin contraction. Two splice variants of long MLCK are expressed in epithelial cells and divergently regulate gut barrier functions; reduced MLCK levels in human colorectal cancers (CRC) with unclarified significance have been reported. CRC are solid tumors clonally sustained by stem cells highly expressing CD44 and CD133. The aim was to investigate the role of MLCK splice variants in CRC tumorigenesis. We found lower MLCK1/2 and higher CD44 expression in human CRC, but no change in CD133 or LGR5. Large-scale bioinformatics showed an inverse relationship between MYLK and CD44 in human sample gene datasets. A 3-fold increased tumor burden was observed in MLCK(-/-) mice compared with wild-type (WT) mice in a chemical-induced CRC model. Primary tumorspheres derived from the MLCK(-/-) mice displayed larger sizes and higher CD44 transcript levels than those from the WT mice. Bioinformatics revealed binding of TEAD4 (a transcriptional enhancer factor family member in the Hippo pathway) to CD44 promoter, which was confirmed by luciferase reporter assay. Individually expressing MLCK1 and MLCK2 variants in the MLCK-knockout (KO) Caco-2 cells inhibited the nuclear localization of TEAD4 cofactors, VGLL3 and YAP1, respectively, and both variants reduced the CD44 transcription. Accelerated cell cycle transit was observed in the MLCK-KO cells, whereby expression of MLCK1/2 variants counterbalanced the cell hyperproliferation. In conclusion, MLCK1/2 variants are novel tumor suppressors by downregulating the TEAD4/CD44 axis via reducing nuclear translocation of distinct transcriptional coactivators. The reduction of epithelial MLCKs, especially isoform 2, may drive cancer stemness and tumorigenesis.

"Metalloestrogenic" effects of cadmium downstream of G protein-coupled estrogen receptor and mitogen-activated protein kinase pathways in human uterine fibroid cells.

Cadmium (Cd) is a toxic metal reported to act as an estrogen "mimic" in the rat uterus and in vitro. We have reported that Cd stimulates proliferation of estrogen-responsive human uterine leiomyoma (ht-UtLM; fibroid) cells through nongenomic signaling involving the G protein-coupled estrogen receptor (GPER), with activation of epidermal growth factor receptor (EGFR) and mitogen-activated protein kinase (pMAPK44/42). In this study, we explored Cd-induced mechanisms downstream of MAPK and whether Cd could stimulate phosphorylation of Histone H3 at serine 10 (H3Ser10ph) through activated Aurora B kinase (pAurora B), a kinase important in activation of histone H3 at serine 10 during mitosis, and if this occurs via Fork head box M1 (FOXM1) and cyclin D1 immediately downstream of MAPK. We found that Cd increased proliferating cell nuclear antigen (PCNA) and H3Ser10ph expression by immunofluorescence, and that H3ser10ph and pAurora B were coexpressed along the metaphase plate in ht-UtLM cells. In addition, Cd-exposed cells showed higher expression of pMAPK44/42, FOXM1, pAurora B, H3ser10ph, and Cyclin D1 by western blotting. Immunoprecipitation and proximity ligation assays further indicated an association between FOXM1 and Cyclin D1 in Cd-exposed cells. These effects were attenuated by MAPK kinase (MEK1/2) inhibitor. In summary, Cd-induced proliferation of ht-UtLM cells occurred through activation of Histone H3 and Aurora B via FOXM1/Cyclin D1 interactions downstream of MAPK. This provides a molecular mechanism of how Cd acts as an "estrogen mimic" resulting in mitosis in hormonally responsive cells.

Targeting Colorectal Cancer with Conjugates of a Glucose Transporter Inhibitor and 5-Fluorouracil.

Overexpression of glucose transporters (GLUTs) in colorectal cancer cells is associated with 5-fluorouracil (1, 5-FU) resistance and poor clinical outcomes. We designed and synthesized a novel GLUT-targeting drug conjugate, triggered by glutathione in the tumor microenvironment, that releases 5-FU and GLUTs inhibitor (phlorizin (2) and phloretin (3)). Using an orthotopic colorectal cancer mice model, we showed that the conjugate exhibited better antitumor efficacy than 5-FU, with much lower exposure of 5-FU during treatment and without significant side effects. Our study establishes a GLUT-targeting theranostic incorporating a disulfide linker between the targeting module and cytotoxic payload as a potential antitumor therapy.

Prevention of dextran sulfate sodium-induced mouse colitis by the fungal protein Ling Zhi-8 via promoting the barrier function of intestinal epithelial cells.

The fungal immunomodulatory protein Ling Zhi-8 (LZ-8) isolated from Ganoderma lucidum (GL) regulates immune cells and inhibits tumor growth; however, the role of LZ-8 in protecting intestinal epithelial cells (IECs) is unknown. In this study, we aim to investigate the functional effect of LZ-8 on IECs. LZ-8 effectively rescued the pro-inflammatory cytokine-induced loss of tight junctions (TJs) by enhancing transepithelial electrical resistance (TEER), reducing permeability, and maintaining the distribution of TJ proteins, in Caco-2 cells. Mechanistically, LZ-8 blocked the upregulation of myosin light chain kinase (MLCK) and NF-kB activation by TLR2-mediated suppression of cytokine signaling (SOCS)-1 expression. Furthermore, LZ-8 pre-treatment reduced the pathological scores of dextran sulfate sodium (DSS)-induced colitis in mice. These results indicated that LZ-8 protected the barrier function of IECs against inflammation. Thus, LZ-8 may potentially be a novel candidate for treating inflammatory bowel disease (IBD).

Differential Receptor Tyrosine Kinase Phosphorylation in the Uterus of Rats Following Developmental Exposure to Tetrabromobisphenol A.

Tetrabromobisphenol A (TBBPA) is a brominated flame retardant that induces endometrial adenocarcinoma and other uterine tumors in Wistar Han rats; however, early molecular events or biomarkers of TBBPA exposure remain unknown. We investigated the effects of TBBPA on growth factor receptor activation (phospho-RTK) in uteri of rats following early-life exposures. Pregnant Wistar Han rats were exposed to TBBPA (0, 0.1, 25, 250 mg/kg/day) via oral gavage on gestation day 6 through weaning of pups (PND 21). Pups were exposed in utero, through lactation, and by daily gavage from PND 22 to PND 90. Uterine horns were collected (at PND 21, PND 33, PND 90) and formalin-fixed or frozen for histologic, immunohistochemical, phospho-RTK arrays, or western blot analysis. At PND 21, the phosphor-RTKs, FGFR2, FGFR3, TRKC and EPHA1 were significantly increased at different treatment concentrations. Several phospho-RTKs were also significantly overexpressed at PND 33 which included epithelial growth factor receptor (EGFR), Fibroblast Growth Factor Receptor 3-4 (FGFR2, FGFR3, FGFR4), insulin-like growth factor receptor 1 (IGF1R), INSR, AXL, MERTK, PDGFRa and b, RET, Tyrosine Kinase with Immunoglobulin Like and EGF Like Domains 1 and 2 (TIE1; TIE2), TRKA, VEGFR2 and 3, and EPHA1 at different dose treatments. EGFR, an RTK overexpressed in endometrial cancer in women, remained significantly increased for all treatment groups at PND 90. Erb-B2 Receptor Tyrosine Kinase 2 (ERBB2) and IGF1R were overexpressed at PND 33 and remained increased through PND 90, although ERBB2 was statistically significant at PND 90. The phospho-RTKs, FGFR3, AXL, DTK, HGFR, TRKC, VEGFR1 and EPHB2 and 4 were also statistically significant at PND 90 at different dose treatments. The downstream effector, phospho-MAPK44/42 was also increased in uteri of treated rats. Our findings show RTKs are dysregulated following early life TBBPA exposures and their sustained activation may contribute to TBBPA-induced uterine tumors observed in rats later in life.