Development of engineered IL-36γ-hypersecreting Lactococcus lactis to improve the intestinal environment.

Interleukin (IL) 36 is a member of the IL-1-like proinflammatory cytokine family that has a protective role in mucosal immunity. We hypothesized that mucosal delivery of IL-36γ to the intestine would be a very effective way to prevent intestinal diseases. Here, we genetically engineered a lactic acid bacterium, Lactococcus lactis, to produce recombinant mouse IL-36γ (rmIL-36γ). Western blotting and enzyme-linked immunosorbent assay results showed that the engineered strain (NZ-IL36γ) produced and hypersecreted the designed rmIL-36γ in the presence of nisin, which induces the expression of the recombinant gene. We administered NZ-IL36γ to mice via oral gavage, and collected the ruminal contents and rectal tissues. Colony PCR using primers specific for NZ-IL36γ, and enzyme-linked immunosorbent assay to measure the rmIL-36γ concentrations of the ruminal contents showed that NZ-IL36γ colonized the mouse intestines and secreted rmIL-36γ. A microbiota analysis revealed increased abundances of bacteria of the genera Acetatifactor, Eubacterium, Monoglobus, and Roseburia in the mouse intestines. Real-time quantitative PCR of the whole colon showed increased Muc2 expression. An in vitro assay using murine colorectal epithelial cells and human colonic cells showed that purified rmIL-36γ promoted Muc2 gene expression. Taken together, these data suggest that NZ-IL36γ may be an effective and attractive tool for delivering rmIL-36γ to improve the intestinal environment.

Dextran-based nanoparticles with 5-FU-conjugated polymethacrylate segments for drug delivery: Synthesis of amphiphilic graft copolymers by mechanochemical solid-state polymerization and characterization.

Dextran (Dx) is a biodegradable and biocompatible polysaccharide, thus promising as a drug delivery carrier for tumor therapy. Herein, we applied mechanical energy to a high molecular weight Dx to control its molecular weight and simultaneously generate mechanoradicals. The solid-state polymerization of methacrylate- or methacrylamide derivatives initiated with Dx mechanoradicals showed polymer conversion of >95%, yielding Dx-based graft copolymers with molecular weights of approximately 30,000 g mol-1. The Dx-based graft copolymers with hydrophobic segments formed nanoparticles with a particle size of 25-35 nm in an aqueous solution. The anti-pancreatic tumor drug 5-fluorouracil (5-FU) was covalently conjugated onto the hydrophobic segments of the amphiphilic Dx, and the nanoparticles were also prepared. The drug release profile from 5-FU-conjugated nanoparticles corresponded well to the Korsmeyer-Peppas model applied to drug release from matrix substrates, and was also immensely predicted by the Logistic and Gompertz curves. The 5-FU-conjugated nanoparticles showed cytotoxicity against the pancreatic adenocarcinoma cell lines (BxPC-3) that were not significantly inferior to the 5-FU positive group. Furthermore, the fluorescein-labeled nanoparticles internalized into BxPC-3 within 6 h and actively migrated into the cytosol. These results suggest that Dx-based graft copolymers with hydrophobic segments might be used to enhance therapeutic activity.

Case Report: A Very Low Birth Weight Female Infant With Congenital Bilateral Periventricular Leukomalacia, Born to a Mother With Coronavirus Disease 2019.

A 26-year-old primipara woman with COVID-19 performed an emergency Cesarean section due to further hypoxemia at 28 weeks 5/7 days gestation. The female neonate was born weighing 1,347 gram with an Apgar score of four at 1 min, three at 5 min, and eight at 10 min. RT-PCR from nasopharyngeal swabs for COVID-19 were performed at birth, 24 h, and 48 h after birth, all of which were negative. On head ultrasound bilateral cystic lesions were found in the anterior horn of the lateral ventricles at birth. A brain magnetic resonance imaging (MRI) test at 56 days of life (corrected 36 weeks and 6/7 days) revealed cystic lesions with T1 low signal, T2 high signal, and T2 Flair high signal around the anterior horn of the lateral ventricle and We diagnose it as Grade 2 periventricular leukomalacia (PVL). She was discharged on day 64 of life, with no abnormality on exam. While the majority of neonates born to women with COVID-19 during pregnancy have favorable outcome, we report a case of a neonate with Grade 2 periventricular leukomalacia and this should prompt clinicians to monitor fetal cerebral function and structure shortly after birth.

Protein-protein interactions between jasmonate-related master regulator MYC and transcriptional mediator MED25 depend on a short binding domain.

A network of protein-protein interactions (PPI) is involved in the activation of (+)-7-iso-jasmonoyl-L-isoleucine (JA-Ile), a plant hormone that regulates plant defense responses as well as plant growth and development. In the absence of JA-Ile, inhibitory protein jasmonate-ZIM-domain (JAZ) represses JA-related transcription factors, including a master regulator, MYC. In contrast, when JA-Ile accumulates in response to environmental stresses, PPI occurs between JAZ and the F-box protein COI1, which triggers JAZ degradation, resulting in derepressed MYC that can interact with the transcriptional mediator MED25 and upregulate JA-Ile-related gene expression. Activated JA signaling is eventually suppressed through the catabolism of JA-Ile and feedback suppression by JAZ splice variants containing a cryptic MYC-interacting domain (CMID). However, the detailed structural basis of some PPIs involved in JA-Ile signaling remains unclear. Herein, we analyzed PPI between MYC3 and MED25, focusing on the key interactions that activate the JA-Ile signaling pathway. Biochemical assays revealed that a short binding domain of MED25 (CMIDM) is responsible for the interaction with MYC, and that a bipartite interaction is critical for the formation of a stable complex. We also show the mode of interaction between MED25 and MYC is closely related to that of CMID and MYC. In addition, quantitative analyses on the binding of MYC3-JAZs and MYC3-MED25 revealed the order of binding affinity as JAZJas < MED25CMIDM < JAZCMID, suggesting a mechanism for how the transcriptional machinery causes activation and negative feedback regulation during jasmonate signaling. These results further illuminate the transcriptional machinery responsible for JA-Ile signaling.

PET/MRI is useful for early detection of pelvic insufficiency fractures after radiotherapy for cervical cancer.

Radiotherapy (RT) is used to manage cervical cancer, and pelvic insufficiency fracture (PIF) is known as a late complication of RT. The present study identified risk factors for PIF after radiotherapy for cervical cancer, and investigated its incidence rate. It also considered the usefulness of positron emission tomography/magnetic resonance imaging (PET/MRI) in PIF diagnosis. A total of 149 patients with cervical cancer who received definitive or adjuvant RT with/without concurrent chemotherapy between January 2013 and December 2018 were investigated in the present study and followed up for more than one month after RT at Kobe University Hospital. The median follow-up period was 32 months (range, 1-87 months), and the median age of all patients was 66 years (age range, 34-90 years). Computed tomography (CT), MRI, PET/CT or PET/MRI were used for image examination. Among the 149 patients, 31 (20.8%) developed PIF. The median age of these patients was 69 years (age range, 44-87 years). Univariate analysis using the log-rank test demonstrated that age (≥60 years) was significantly associated with PIF. The median maximum standardized uptake value of PIF sites on PET/CT was 4.32 (range, 3.04-4.81), and that on PET/MRI was 3.97 (range, 1.21-5.96) (P=0.162). Notably, the detection time of PIF by PET/MRI was significantly earlier compared with PET/CT (P<0.05). The incidence of PIF after RT for cervical cancer was 20.8%, and age was significantly associated with risk factors for such fractures. Taken together, these results suggest that PET/MRI, which offers the advantage of decreased radiation exposure to the patient, is useful for diagnosing PIF and can detect it earlier than PET/CT imaging.

Correction: Rational design of a stapled JAZ9 peptide inhibiting protein-protein interaction of a plant transcription factor.

[This corrects the article DOI: 10.1039/D0CB00204F.].

Rational design of a stapled JAZ9 peptide inhibiting protein-protein interaction of a plant transcription factor.

We herein describe the development of a stapled peptide inhibitor for a jasmonate-related transcription factor. The designed peptide selectively inhibited MYCs, master-regulators of jasmonate signaling, and selectively suppressed MYC-mediated gene expression in Arabidopsis thaliana. It is proposed as a novel chemical tool for the analysis of MYC related jasmoante signaling.

Genetic Variation in Y-Chromosome Genes of Sika Deer (Cervus nippon) in Japan.

Sika deer (Cervus nippon) in Japan are classified into southern and northern groups. However, previous studies primarily relied on maternally inherited mitochondrial DNA (mtDNA). The paternally inherited Y-chromosome is useful for analyzing the contribution of males to the population genetic history of sika deer. In total, approximately 16 kb of partial sequences of four Y-chromosomal genes, Y-linked, sex-determining region Y, DEAD-box helicase 3 Y-linked, and Zinc finger protein Y-linked, were sequenced to investigate intraspecific variation. As a result, we identified nine intronic single nucleotide polymorphisms (SNPs) in 478 sika deer samples collected over the entire Japanese archipelago from Hokkaido to Kyushu. SNP genotyping revealed 10 distinct haplotypes (SYH1-SYH10). The most common haplotype (SYH1) was present in all populations and was the most abundant haplotype, identified in 80.3% of the sampled individuals. The remaining haplotypes were unique to a single locality. SYH1 was also central to all other haplotypes that diverged by a SNP, resulting in this haplotype being the core of a star-like cluster topography. We found that contrary to mtDNA patterns, there was no clear differentiation of Y-chromosome markers between the southern and the northern populations. Due to the female philopatry of sika deer, mtDNA may provide a highly structured differentiation of populations. On the other hand, the male-biased gene flow may provide a reduced differentiation of populations. Our findings revealed that the genetic structure of the Japanese sika deer is more complex than previously thought based on mtDNA-based phylogeographic studies.

A Case of Significant Response to Olaparib in a Patient with Primary Peritoneal Carcinosarcoma Diagnosed by Laparoscopic Surgery.

Primary peritoneal carcinosarcomas which arise from extragenital locations are extremely rare. Carinosarcomas contain both carcinomatous and sarcomatous elements and can be mainly detected in the female genital tract. We herein report a case of primary peritoneal carcinosarcoma diagnosed by laparoscopic surgery and treated with olaparib. A 62-year-old woman referred to our hospital due to abdominal distension. From imaging findings, we suspected advanced primary peritoneal carcinoma, and laparoscopic surgery was thereafter performed. The pathological diagnosis was carcinosarcoma, and the patient received chemotherapy with docetaxel and carboplatin. After three cycles of chemotherapy, the interval debulking surgery was attempted but resulted in suboptimal results. Because the bilateral ovaries were observed with a normal size and normal findings, we considered that the most likely diagnosis was primary peritoneal carcinosarcoma. After the additional chemotherapy and a 6-month observation period, the tumor relapsed. The patient received chemotherapy again, and the peritoneal carcinosarcoma was judged to be a platinum-sensitive tumor. Oral administration of olaparib was thus initiated. Although a dose reduction was needed due to anemia, olaparib was effective, and the patient could continue the drug for another 7 months. This is the first report of primary peritoneal carcinosarcoma treated with olaparib and shows that it could be a treatment option for platinum-sensitive tumors.

A Fluorescence Anisotropy-Based Comprehensive Method for the In Vitro Screening of COI1-JAZs Agonists and Antagonists.

The phytohormone (+)-7-iso-jasmonoyl-L-isoleucine (JA-Ile) causes protein-protein interactions (PPI) between F-box Protein CORONATINE INSENSITIVE 1 (COI1) and JASMONATE ZIM DOMAIN (JAZ) transcriptional repressor. A total of 13 JAZ subtypes are encoded in the genome of Arabidopsis thaliana; however, their genetic redundancy obfuscates the individual function of each JAZ. One approach to decipher this redundant signaling network is chemical genetics, using small molecules specific to individual JAZ subtype, for which a reliable and high-throughput screening system of the ligands for all combinations of COI1-JAZs would be indispensable. In this chapter, we describe a fluorescence anisotropy-based quantitative screening system for the ligands of COI1-JAZ co-receptors. Our method is applicable to agonists and antagonists of the COI1-JAZs.

Near-Infrared-Emitting Nitrogen-Doped Nanographenes.

The quantum-size effect, which enables nanographenes to emit photoluminescence (PL) in the UV to visible region, has inspired intense research. However, the control of the PL properties of nanographenes through manipulation of their π-system by post-modifications is not well developed. By utilizing a ring-closure reaction between an aromatic 1,2-dicarboxylic acid and a 1,8-naphthalenediamine derivative, which produces a perimidine framework, nitrogen-doped nanographenes were realized. Two nanographenes produced by a one-pot reaction of edge-oxidized nanographene (GQD-2) with 1,8-naphthalenediamine derivatives (GQD-1 a and GQD-1 b) displayed an absorption band extending to >1000 nm; furthermore, the PL wavelength of GQD-1 a was significantly red-shifted into the near-infrared (NIR) region in which it can be used for bioimaging. Time-dependent DFT calculations of model nanographenes showed that the functional groups narrow the HOMO-LUMO gap, realizing the NIR-emitting nanographenes.

Risk factors for deep venous thrombosis in women with ovarian cancer.

We aim to clarify the incidence of deep venous thrombosis (DVT) before treatment in women with ovarian cancer and identify risk factors for DVT.In this prospective study, 110 women underwent venous ultrasonography before cancer treatment and D-dimer levels were measured. We investigated factors predicting DVT by logistic regression.DVT was detected in 25 of 110 women (22.7%) and pulmonary thromboembolism was coexisted in 2 women (1.8%). A total of 21 women (84.4%) with DVT were asymptomatic. D-dimer levels in women with DVT (median, 10.9; range, <0.5-98.2 µg/mL) were significantly higher than those in women without DVT (2.0; <0.5-60.8 µg/mL; P < .01). When 10.9 µg/mL was used as a cutoff value for D-dimer levels to predict DVT, specificity, sensitivity, and positive and negative predictive values were 92.9%, 52.0%, 68.4%, and 86.8%, respectively. The multivariate analysis demonstrated that D-dimer level (odds ratio [OR], 19.7; 95% confidence interval [CI], 5.89-76.76) and clear cell histology (OR, 7.1; 95% CI, 2.12-25.67) were independent factors predicting DVT.Asymptomatic DVT occurred with great frequency before treatment in patients with ovarian cancer. High D-dimer level and clear cell pathology is associated with a higher DVT risk.

Photoluminescence responses of graphene quantum dots toward organic bases and an acid.

Edge-modified graphene quantum dots (GQD-1) showed base-dependent photoluminescence (PL) responses. DBN, DBU, and Et3N gradually enhanced the PL intensity and then caused quenching, whereas quenching was not observed when pyridine and pyrimidine were used. The quenching can be explained by the photoinduced electron transfer from the anchored bases at the periphery to the emissive sites of GQD-1.

Relaxin has anti-apoptotic effects on human trophoblast-derived HTR-8/SV neo cells.

The study was conducted to evaluate the effects of human relaxin on apoptosis in the human trophoblast derived HTR-8/SV neo cell line, which is a possible model of human extravillous trophoblasts (EVTs). HTR-8/SV neo cells, cultured in phenol red free RPMI1640 medium, were treated with different doses of human recombinant (rH2) relaxin in serum-deprived conditions. RT-PCR was used for evaluating relaxin receptor: RXFP1 and RXFP2 expression in HTR-8/SV neo cells. The cell death was examined by TUNEL assay. Furthermore, we investigated caspase-3, cleaved PARP and Bcl-2 expressions by Western blot analysis to recognize the translational effects of anti-apoptotic and pro-apoptotic proteins. RXFP1 and RXFP2 mRNA expression was observed in HTR-8/SV neo cells. Compared with untreated control cultures, treatment with rH2 relaxin, decreased TUNEL-positive rate in HTR-8/SV neo cells was observed. Western blot analysis revealed that treatment with rH2 relaxin decreased the expression of caspase-3 and cleaved PARP, but in contrast increased Bcl-2 expression in those cells. These results suggest that rH2 relaxin has anti-apoptotic effects on HTR8/SV neo cells by decreasing pro-apoptotic caspase-3 and cleaved PARP expression and up-regulating anti-apoptotic Bcl-2 expression.

Recombinant H2 relaxin inhibits apoptosis and induces cell proliferation in cultured leiomyoma cells without affecting those in cultured normal myometrial cells.

OBJECTIVE: To evaluate the effects of human relaxin on proliferation and apoptosis in cultured human uterine leiomyoma cells and normal myometrial cells. DESIGN: In vitro experiment. SETTING: Research laboratory at Kobe University Graduate School of Medicine. PATIENT(S): Nine patients undergoing hysterectomy for uterine leiomyoma. INTERVENTION(S): Cultured leiomyoma cells and normal myometrial cells were treated with human recombinant (rH2) relaxin. MAIN OUTCOME MEASURE(S): Human relaxin receptor LGR7 expressions in cultured leiomyoma cells and myometrial cells were evaluated by immunocytochemical staining. Cell proliferation, proliferating cell nuclear antigen-positive rate, and TUNEL-positive rate were assessed by MTS assay, immunocytochemistry, and TUNEL assay, respectively. Caspase-3 expression was evaluated by Western blot analysis. RESULT(S): LGR7 expression was observed both in cultured human leiomyoma cells and myometrial cells. Compared with untreated control cultures, treatment with rH2 relaxin increased the number of viable cultured leiomyoma cells and the proliferating cell nuclear antigen-positive rate in those cells but not in myometrial cells. Moreover, treatment with rH2 relaxin decreased the TUNEL-positive rate in cultured leiomyoma cells but not in myometrial cells. Similarly, Western blot analysis revealed that treatment with rH2 relaxin decreased the expression of caspase-3 in cultured leiomyoma cells but not in myometrial cells. CONCLUSION(S): These results suggest that rH2 relaxin selectively inhibits apoptosis by down-regulating caspse-3 expression and induces proliferation in cultured human leiomyoma cells without affecting apoptosis or proliferation in normal myometrial cells.

The lowest surviving birth weight infant delivered from a systemic lupus erythematosus mother with antiphospholipid syndrome.

We report the intact surviving case of a newborn with a birth weight of 412 g delivered from an active systemic lupus erythematosus (SLE) mother with antiphospholipid syndrome. A review of the literature revealed that our infant is the lowest surviving birth weight in newborns from SLE mothers to date.