Sex differences in colonic gene expression and fecal microbiota composition in a mouse model of obesity-associated colorectal cancer.

This study investigated the sex-specific correlation between obesity and colorectal cancer emphasizing a more pronounced association in males. Estrogen, chromosomal genes, and gut bacteria were assessed in C57BL6/J male, female and ovariectomized (OVX) female mice, subjected to either a low-fat diet (LFD) or high-fat diet (HFD) for 14 weeks. Induction of colon tumor involved azoxymethane (10 mg/kg) administration, followed by three cycles of dextran sulfate sodium. Male mice on HFD exhibited higher final body weight and increased colon tumors compared to females. Colonic mucin 2 expression was significantly higher in females. HFD-modulated differentially expressed genes numbered 290 for males, 64 for females, and 137 for OVX females. Only one up-regulated gene (Gfra3) overlapped between females and OVX females, while two down-regulated genes (Thrsp and Gbp11) overlapped between males and OVX females. Genes up-regulated by HFD in males were linked to cytokine-cytokine interaction, HIF-1 signaling pathway, central carbon metabolism in cancer. Sex-specific changes in gut microbial composition in response to HFD were observed. These findings suggest a male-specific vulnerability to HFD-induced colon tumor formation, implicating key genes and colonic bacteria in colon tumorigenesis.

The therapeutic effects of vitamin D3 administration on the embryo implantation.

Various adjuvants have been tested clinically for patients with problems with embryo implantation during in vitro fertilization (IVF)-embryo transfer (ET). Vitamin D3, an essential modulator of various physiological processes, has received attention as an important adjuvant for successful pregnancy, as many studies have shown a strong association between vitamin D deficiency and implantation failure and fetal growth restriction. However, vitamin D has been widely utilized in different protocols, resulting in non-reproducible and debatable outcomes. In the present study, we demonstrated that cyclic intrauterine administration of vitamin D3 increased endometrial receptivity and angiogenesis, which could be attributed to increased recruitment of uterus-resident natural killer cells. In particular, cyclic treatment of vitamin D3 promoted stable attachment of the embryo onto endometrial cells in vitro, suggesting its merit during the early stage of embryo implantation to support the initial maternal-fetal interactions. Our findings suggest that women with repeated implantation failure may benefit from the use of vitamin D3 as a risk-free adjuvant prior to IVF-ET procedures to improve the uterine environment, and make it favorable for embryo implantation.

Endometrial organoids: a reservoir of functional mitochondria for uterine repair.

Background: Asherman's syndrome (AS) is a dreadful gynecological disorder of the uterus characterized by intrauterine adhesion with severe fibrotic lesions, resulting in a damaged basalis layer with infertility. Despite extensive research on overcoming AS, evidence-based effective and reproducible treatments to improve the structural and functional morphology of the AS endometrium have not been established. Methods: Endometrial organoids generated from human or mouse endometrial tissues were transplanted into the uterine cavity of a murine model of AS to evaluate their transplantable feasibility to improve the AS uterine environment. The successful engraftment of organoid was confirmed by detection of human mitochondria and cytosol (for human endometrial organoid) or enhanced green fluorescent protein signals (for mouse endometrial organoid) in the recipient endometrium. The therapeutic effects mediated by organoid transplantation were examined by the measurements of fibrotic lesions, endometrial receptivity and angiogenesis, and fertility assessment by recording the number of implantation sites and weighing the fetuses and placenta. To explore the cellular and molecular mechanisms underlying the recovery of AS endometrium, we evaluated the status of mitochondrial movement and biogenetics in organoid transplanted endometrium. Results: Successfully engrafted endometrial organoids with similar morphological and molecular features to the parental tissues dramatically repaired the AS-induced damaged endometrium, significantly reducing fibrotic lesions and increasing fertility outcomes in mice. Moreover, dysfunctional mitochondria in damaged tissues, which we propose might be a key cellular feature of the AS endometrium, was fully recovered by functional mitochondria transferred from engrafted endometrial organoids. Endometrial organoid-originating mitochondria restored excessive collagen accumulation in fibrotic lesions and shifted uterine metabolic environment to levels observed in the normal endometrium. Conclusions: Our findings suggest that endometrial organoid-originating mitochondria might be key players to mediate uterine repair resulting in fertility enhancement by recovering abrogated metabolic circumstance of the endometrium with AS. Further studies addressing the clinical applicability of endometrial organoids may aid in identifying new therapeutic strategies for infertility in patients with AS.

The therapeutic effects and optimal timing of granulocyte colony stimulating factor intrauterine administration during IVF-ET.

Most of embryos fail to produce live offspring during In Vitro Fertilization-Embryo Transfer (IVF-ET) procedure. There is a dearth of research activity addressing this problem despite the significant population of women suffering from repeated implantation failure after transfer of high-quality of embryos. As a clinically accessible option, granulocyte colony stimulating factor (G-CSF) is often used for the treatment to improve the rates of embryo implantation. However, there are currently no evidence-based standardized protocol for the clinical use of G-CSF. G-CSF was administered into one side of mouse uterine horns and saline was infused into the other side of horns as a control. Intrauterine G-CSF administration showed maximal effects 24 h after administration in enhancing endometrial receptivity and subsequent increase of angiogenesis by demonstrating elevated integrin ß3 and OPN and reduced cytotoxicity of NK cells. Furthermore, G-CSF administration 24 h prior to embryo transfer promoted the stability of attached embryos at the early stage of implantation in vitro. Our findings suggest as new consensus criteria providing a potential therapeutic strategy of the clinical use of G-CSF to achieve maximal effects of IVF-ET for patients who are suffering from repeated implantation failure with the problems with endometrial receptivity.

Mobile Efficient Diagnostics of Infectious Diseases via On-Chip RT-qPCR: MEDIC-PCR.

The COVID-19 outbreak has caused public and global health crises. However, the lack of on-site fast, reliable, sensitive, and low-cost reverse transcription polymerase chain reaction (RT-PCR) testing limits early detection, timely isolation, and epidemic prevention and control. Here, the authors report a rapid mobile efficient diagnostics of infectious diseases via on-chip -RT-quantitative PCR (RT-qPCR): MEDIC-PCR. First, the authors use a roll-to-roll printing process to accomplish low-cost carbon-black-based disposable PCR chips that enable rapid LED-induced photothermal PCR cycles. The MEDIC-PCR can perform RT (3 min), and PCR (9 min) steps. Further, the cohort of 89 COVID-19 and 103 non-COVID-19 patients testing is completed by the MEDIC-PCR to show excellent diagnostic accuracy of 97%, sensitivity of 94%, and specificity of 98%. This MEDIC-PCR can contribute to the preventive global health in the face of a future pandemic.

High-Throughput Screening of Anti-cancer Drugs Using a Microfluidic Spheroid Culture Device with a Concentration Gradient Generator.

Tumor spheroid models are widely used for drug screening as in vitro models of the tumor microenvironment. There are various ways in which tumor spheroid models can be prepared, including the self-assembly of cells using low-adherent plates, micro-patterned plates, or hanging-drop plates. Recently, drug high-throughput screening (HTS) approaches have incorporated the use of these culture systems. These HTS culture systems, however, require complicated equipment, such as robot arms, detectors, and software for handling solutions and data processing. Here, we describe protocols that allow tumor spheroids to be tested with different concentrations of a drug in a parallel fashion using a microfluidic device that generates a gradient of anti-cancer drugs. This microfluidic spheroid culture device with a concentration gradient generator (µFSCD-CGG) enables the formation of 50 tumor spheroids and the testing of drugs at five different concentrations. First, we provide a protocol for the fabrication of the µFSCD-CGG, which has both a culture array in which tumor cells are injected and aggregate to form spheroids and a concentration gradient generator for drug testing. Second, we provide a protocol for tumor spheroid formation and HTS of anti-cancer drugs using the device. Finally, we provide a protocol for assessing the response of tumor spheroids at different drug concentrations. To address the needs of the pharmaceutical industry, this protocol can be used for various cell types, including stem cells, and the number of tumor spheroids and drug concentration ranges that can be tested in the µFSCD-CGG can be increased. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Fabrication of a microfluidic spheroid culture device with a concentration gradient generator (µFSCD-CGG) Basic Protocol 2: Seeding cells and formation of spheroids in the µFSCD-CGG Basic Protocol 3: Drug treatment and assessment of cell viability in the µFSCD-CGG.

Gut-Kidney Axis on Chip for Studying Effects of Antibiotics on Risk of Hemolytic Uremic Syndrome by Shiga Toxin-Producing Escherichia coli.

Shiga toxin-producing Escherichia coli (STEC) infects humans by colonizing the large intestine, and causes kidney damage by secreting Shiga toxins (Stxs). The increased secretion of Shiga toxin 2 (Stx2) by some antibiotics, such as ciprofloxacin (CIP), increases the risk of hemolytic-uremic syndrome (HUS), which can be life-threatening. However, previous studies evaluating this relationship have been conflicting, owing to the low frequency of EHEC infection, very small number of patients, and lack of an appropriate animal model. In this study, we developed gut-kidney axis (GKA) on chip for co-culturing gut (Caco-2) and kidney (HKC-8) cells, and observed both STEC O157:H7 (O157) infection and Stx intoxication in the gut and kidney cells on the chip, respectively. Without any antibiotic treatment, O157 killed both gut and kidney cells in GKA on the chip. CIP treatment reduced O157 infection in the gut cells, but increased Stx2-induced damage in the kidney cells, whereas the gentamycin treatment reduced both O157 infection in the gut cells and Stx2-induced damage in the kidney cells. This is the first report to recapitulate a clinically relevant situation, i.e., that CIP treatment causes more damage than gentamicin treatment. These results suggest that GKA on chip is very useful for simultaneous observation of O157 infections and Stx2 poisoning in gut and kidney cells, making it suitable for studying the effects of antibiotics on the risk of HUS.