Efficient intracellular drug delivery by co-administration of two antibodies against cell adhesion molecule 1.

Cell adhesion molecule 1 (CADM1), a single-pass transmembrane protein, is involved in oncogenesis. We previously demonstrated the therapeutic efficacy of anti-CADM1 ectodomain monoclonal antibodies against mesothelioma; however, the underlying mechanism is unclear. In the present study, we explored the molecular behavior of anti-CADM1 antibodies in CADM1-expressing tumor cells. Sequencing analyses revealed that the anti-CADM1 chicken monoclonal antibodies 3E1 and 9D2 are IgY and IgM isotype antibodies, respectively. Co-administration of 3E1 and 9D2 altered the subcellular distribution of CADM1 from the detergent-soluble fraction to the detergent-resistant fraction in tumor cells. Using recombinant chicken-mouse chimeric antibodies that had been isotype-switched from IgG to IgM, we demonstrated that the combination of the variable region of 3E1 and the constant region of IgM was required for CADM1 relocation. Cytochemical studies showed that 3E1 colocalized with late endosomes/lysosomes after co-administration with 9D2, suggesting that the CADM1-antibody complex is internalized from the cell surface to intracellular compartments by lipid-raft mediated endocytosis. Finally, 3E1 was conjugated with the antimitotic agent monomethyl auristatin E (MMAE) via a cathepsin-cleavable linker. Co-administration of 3E1-monomethyl auristatin E and 9D2 suppressed the growth of multiple types of tumor cells, and this anti-tumor activity was confirmed in a syngeneic mouse model of melanoma. 3E1 and 9D2 are promising drug delivery vehicles for CADM1-expressing tumor cells.

[A case of primary central nervous system lymphoma of the sellar region presented with panhypopituitarism].

The patient is a 41-year-old woman. She presented with vomiting and lightheadedness, and blood tests showed a generalized decrease in pituitary hormones and hyperprolactinemia. A head MRI showed increased signal intensity lesions on FLAIR image in the pituitary stalk, corpus callosum, periventricular area of the fourth ventricle, and superior cerebellar peduncle. The lesions were homogeneously enhanced, and a brain biopsy confirmed the diagnosis of primary diffuse large B-cell lymphoma of the central nervous system, and chemotherapy was started. Although the suprasellar region is a rare site for primary central nervous system lymphoma (PCNSL), it should be diagnosed early by biopsy.

Long-term endoscopic change of gastric polyp associated with administration of vonoprazan.

Vonoprazan (VPZ) has been available in Japan since 2015. Endoscopic features of proton-pump inhibitor (PPI)-related gastric mucosal changes, including fundic gland and hyperplastic polyps, have been observed. However, the relationship between gastric polyps and VPZ remains unclear. A 65-year-old man with reflux esophagitis-associated symptoms refractory to PPI was referred to our hospital. VPZ (20 mg) was administered for 3 weeks, which proved effective. Afterward, VPZ dose was reduced to 10 mg; the reflux symptoms worsened, and 20 mg VPZ was restarted. Afterward, esophagogastroduodenoscopy (EGD) revealed a gradually enlarging gastric polyp in the cardia. After 5 years of VPZ administration, the patient developed a reddish polyp (approximately 10 mm) with a whitish substance in the cardia. Based on the clinical course, the polyp was considered to have enlarged because of the long-term VPZ administration. After being informed of the endoscopic findings, the patient decided to discontinue VPZ. One year after VPZ discontinuation, EGD revealed a shrunken polyp (5 mm). Long-term acid suppression causes hypergastrinemia, which may lead to gastric mucosal changes, including gastric polyps. There are few case reports of a decrease in the number and size of gastric polyps after VPZ discontinuation. Hence, some VPZ-induced endoscopic changes may be reversible.

Hyposecretion of cervical MUC5B is related to preterm birth in pregnant women after cervical excisional surgery.

PROBLEM: Excisional surgery for cervical intraepithelial neoplasia is a risk factor for preterm birth in subsequent pregnancies. However, the underlying mechanisms of this association remain unclear. We previously showed that cervical MUC5B, a mucin protein, may be a barrier to ascending pathogens during pregnancy. We thus hypothesized that hyposecretion of cervical MUC5B is associated with preterm birth after cervical excisional surgery. METHOD OF STUDY: This prospective nested case-control study (Study 1) included pregnant women who had previously undergone cervical excisional surgery across 11 hospitals. We used proteomics to compare cervicovaginal fluid at 18-22 weeks of gestation between the preterm and term birth groups. In another case-control analysis (Study 2), we compared MUC5B expression in nonpregnant uterine tissues between 15 women with a history of cervical excisional surgery and 26 women without a history of cervical surgery. RESULTS: The abundance of MUC5B in cervicovaginal fluid was significantly decreased in the preterm birth group (fold change = 0.41, p = .035). Among the 480 quantified proteins, MUC5B had the second highest positive correlation with gestational age at delivery in the combined preterm and term groups. The cervicovaginal microbiome composition was not significantly different between the two groups. Cervical length was not correlated with gestational age at delivery (r = 0.18, p = .079). Histologically, the MUC5B-positive area in the nonpregnant cervix was significantly decreased in women with a history of cervical excisional surgery (0.85-fold, p = .048). The distribution of MUC5B-positive areas in the cervical tissues of 26 women without a history of cervical excisional surgery differed across individuals. CONCLUSIONS: This study suggests that the primary mechanism by which cervical excisional surgery causes preterm birth is the hyposecretion of MUC5B due to loss of the cervical glands.

Metastatic Sites in Rare Genitourinary Malignancies and Primary Cancer Sites in Genitourinary Organ Metastases: A Secondary Analysis Using the Japanese Pathological Autopsy Registry Database.

Background: The epidemiology of metastases from rare genitourinary cancer and metastases to genitourinary organs from other primary neoplasms remains poorly understood. Objective: To investigate the epidemiology of rare genitourinary metastases from rare genitourinary organ-type cancer and to genitourinary organs using data from a large national autopsy registry in Japan. Design setting and participants: A secondary analysis of the data reported in the Annual of the Pathological Autopsy Cases in Japan and the Japanese Mortality Database from 1993 to 2020 was performed. Outcome measurements and statistical analysis: Via a retrospective epidemiologic analysis, we evaluated the frequency (probability of occurrence [number per person]) and proportion (percentage) of metastases from upper urinary tract, adrenal, testicular, urethral, and penile cancers. Moreover, the sites of primary tumors metastasizing to genitourinary organs were examined. Results and limitations: In Japan, the mortality rate of upper urinary tract cancer is increasing rapidly. In the integrated database with 365 099 autopsies and 835 959 metastatic organs, the major metastatic sites (range of frequency ratios) of rare genitourinary organ-type cancers were the lungs (0.38-0.47), liver (0.21-0.56), bone (0.16-0.33), adrenal gland (0.10-0.20), peritoneum (0.0-0.16), and kidneys (0.07-0.22). The major primary sites (range of proportions) of genitourinary organ metastases were the respiratory tract (5.6-34.0%), stomach (4.7-27.0%), hematologic site (0.9-24.9%), lymphoid (2.4-22.2%), bladder (0.8-20.0%), prostate (0.7-14.1%), rectal (2.0-11.7%), and pancreas (2.6-11.0%). The cancers with a high likelihood of genitourinary metastasis were respiratory and stomach cancers. However, the study lacked individual-level information, and there might be a concomitant selection bias in this autopsy study. Conclusions: This large-scale autopsy database analysis identified the epidemiology of metastasis from rare genitourinary organ-type cancer and the origins of metastasis to genitourinary organs. Patient summary: This study provides valuable metastatic epidemiologic data and clinical information that are fundamental to the mechanisms of genitourinary metastasis.

Persistent iatrogenic muscular ventricular septal lead perforation after pacemaker implantation using delivery sheath system.

Ventricular septal perforation is a rare complication of pacemaker implantation. Here, we describe the case of a 69-year-old man with complete atrioventricular block and heart failure. The right ventricular pacemaker was implanted with a long pre-shaped delivery sheath. A new systolic murmur appeared after the procedure. Transthoracic echocardiography revealed a ventricular septal perforation, with a Qp/Qs of 1.09, which was a small shunt rate and required no intervention. The persistent ventricular septal perforation was observed, and the shunt rate remained at 8-month follow-up. Learning objective: Ventricular septal lead perforation (VSP) is a rare complication of pacemaker implantation. Although iatrogenic VSP generally close spontaneously without adverse clinical outcomes, clinicians should pay attention to the possibility of its persistence.

Complete response of metastatic papillary renal cell carcinoma with inferior vena cava tumor thrombus to nivolumab plus cabozantinib.

Introduction: The effectiveness of nivolumab plus cabozantinib for metastatic papillary renal cell carcinoma with inferior vena cava tumor thrombus remains unclear. Case presentation: A 77-year-old male was diagnosed with right papillary renal cell carcinoma with a metastatic lesion on Gerota's fascia, lymph node metastasis, and inferior vena cava tumor thrombus. He was treated with nivolumab plus cabozantinib. As all lesions regressed enough to permit complete resection, radical nephrectomy, thrombectomy, and retroperitoneal lymph node dissection were performed. No viable malignant cells were identified histopathologically. Despite the discontinuation of nivolumab plus cabozantinib, there has been no recurrence for 9 months. Conclusion: Nivolumab plus cabozantinib has effectiveness for metastatic papillary renal cell carcinoma with inferior vena cava tumor thrombus.

Rab41-mediated ESCRT machinery repairs membrane rupture by a bacterial toxin in xenophagy.

Xenophagy, a type of selective autophagy, is a bactericidal membrane trafficking that targets cytosolic bacterial pathogens, but the membrane homeostatic system to cope with bacterial infection in xenophagy is not known. Here, we show that the endosomal sorting complexes required for transport (ESCRT) machinery is needed to maintain homeostasis of xenophagolysosomes damaged by a bacterial toxin, which is regulated through the TOM1L2-Rab41 pathway that recruits AAA-ATPase VPS4. We screened Rab GTPases and identified Rab41 as critical for maintaining the acidification of xenophagolysosomes. Confocal microscopy revealed that ESCRT components were recruited to the entire xenophagolysosome, and this recruitment was inhibited by intrabody expression against bacterial cytolysin, indicating that ESCRT targets xenophagolysosomes in response to a bacterial toxin. Rab41 translocates to damaged autophagic membranes via adaptor protein TOM1L2 and recruits VPS4 to complete ESCRT-mediated membrane repair in a unique GTPase-independent manner. Finally, we demonstrate that the TOM1L2-Rab41 pathway-mediated ESCRT is critical for the efficient clearance of bacteria through xenophagy.

Discovery of Selective Histone Deacetylase 1 and 2 Inhibitors: Screening of a Focused Library Constructed by Click Chemistry, Kinetic Binding Analysis, and Biological Evaluation.

Histone deacetylase 1 and 2 (HDAC1/2) inhibitors are potentially useful as tools for probing the biological functions of the isoforms and as therapeutic agents for cancer and neurodegenerative disorders. To discover potent and selective inhibitors, we screened a focused library synthesized by using click chemistry and obtained KPZ560 as an HDAC1/2-selective inhibitor. Kinetic binding analysis revealed that KPZ560 inhibits HDAC2 through a two-step slow-binding mechanism. In cellular assays, KPZ560 induced a dose- and time-dependent increase of histone acetylation and showed potent breast cancer cell growth-inhibitory activity. In addition, gene expression analyses suggested that the two-step slow-binding inhibition by KPZ560 regulated the expression of genes associated with cell proliferation and DNA damage. KPZ560 also induced neurite outgrowth of Neuro-2a cells and an increase in the spine density of granule neuron dendrites of mice. The unique two-step slow-binding character of o-aminoanilides such as KPZ560 makes them interesting candidates as therapeutic agents.

First case report of robot-assisted radical cystectomy for bladder cancer that developed after salvage radiotherapy following radical prostatectomy for prostate cancer.

Introduction: Robot-assisted radical cystectomy for bladder cancer that develops after curative treatment for prostate cancer has not yet been reported. Case presentation: A 65-year-old man underwent radical prostatectomy and received salvage radiotherapy after his postoperative prostate-specific antigen level failed to decrease. Nine years after radiotherapy, local recurrence and lung/bone metastases were observed, and he was started on androgen deprivation therapy. In the following year, he was diagnosed with nonmuscle invasive bladder cancer. He underwent transurethral resection of the bladder tumor once but had multiple recurrences within 3 months. As hematuria could not be controlled by transurethral surgery, he underwent robot-assisted radical cystectomy without rectal injury. Since then, there has been no recurrence of either bladder or prostate cancer. Conclusion: This is the first report of a successful robot-assisted radical cystectomy for bladder cancer that developed after local salvage radiotherapy following radical prostatectomy for prostate cancer.

Proteome analysis of CD5-positive diffuse large B cell lymphoma FFPE tissue reveals downregulation of DDX3X, DNAJB1, and B cell receptor signaling pathway proteins including BTK and Immunoglobulins.

BACKGROUND: The molecular pathology of diffuse large B cell lymphoma (DLBCL) has been extensively studied. Among DLBCL subtypes, the prognosis of CD5-positive DLBCL is worse than that of CD5-negative DLBCL, considering the central nervous system relapse and poor response to R-CHOP therapy. However, the molecular mechanisms underlying the tumorigenesis and progression of CD5-positive DLBCL remain unknown. METHODS: To identify molecular markers that can be targeted for treating DLBCL, a proteomic study was performed using liquid chromatography-mass spectrometry with chemically pretreated formalin-fixed paraffin-embedded specimens from CD5-positive (n = 5) and CD5-negative DLBCL patients (n = 6). RESULTS: Twenty-one proteins showed significant downregulation in CD5-positive DLBCL compared to CD5-negative DLBCL. Principal component analysis of protein expression profiling in CD5-positive and CD5-negative DLBCL revealed that DNAJB1, DDX3X, and BTK, which is one of the B cell phenotypic proteins, were the most significantly downregulated proteins and served as biomarkers that distinguished both groups. Additionally, a set of immunoglobulins, including IgG4, exhibited significant downregulation. Immunohistochemistry analysis for BTK demonstrated reduced staining in CD5-positive DLBCL compared to CD5-negative DLBCL. CONCLUSIONS: In conclusion, DNAJB1 and DDX3X, BTK, and a set of immunoglobulins are promising biomarkers. Probably, the suppression of BCR signaling is the unique phenotype of CD5-positive DLBCL. This formalin-fixed paraffin-embedded (FFPE)-based profiling may help to develop novel therapeutic molecularly targeted drugs for treating DLBCL.

CD44 correlates with longevity and enhances basal ATF6 activity and ER stress resistance.

The naked mole rat (NMR) is the longest-lived rodent, resistant to multiple age-related diseases including neurodegeneration. However, the mechanisms underlying the NMR's resistance to neurodegenerative diseases remain elusive. Here, we isolated oligodendrocyte progenitor cells (OPCs) from NMRs and compared their transcriptome with that of other mammals. Extracellular matrix (ECM) genes best distinguish OPCs of long- and short-lived species. Notably, expression levels of CD44, an ECM-binding protein that has been suggested to contribute to NMR longevity by mediating the effect of hyaluronan (HA), are not only high in OPCs of long-lived species but also positively correlate with longevity in multiple cell types/tissues. We found that CD44 localizes to the endoplasmic reticulum (ER) and enhances basal ATF6 activity. CD44 modifies proteome and membrane properties of the ER and enhances ER stress resistance in a manner dependent on unfolded protein response regulators without the requirement of HA. HA-independent role of CD44 in proteostasis regulation may contribute to mammalian longevity.

Phosphorylation of α-synuclein at T64 results in distinct oligomers and exerts toxicity in models of Parkinson's disease.

α-Synuclein accumulates in Lewy bodies, and this accumulation is a pathological hallmark of Parkinson's disease (PD). Previous studies have indicated a causal role of α-synuclein in the pathogenesis of PD. However, the molecular and cellular mechanisms of α-synuclein toxicity remain elusive. Here, we describe a novel phosphorylation site of α-synuclein at T64 and the detailed characteristics of this post-translational modification. T64 phosphorylation was enhanced in both PD models and human PD brains. T64D phosphomimetic mutation led to distinct oligomer formation, and the structure of the oligomer was similar to that of α-synuclein oligomer with A53T mutation. Such phosphomimetic mutation induced mitochondrial dysfunction, lysosomal disorder, and cell death in cells and neurodegeneration in vivo, indicating a pathogenic role of α-synuclein phosphorylation at T64 in PD.

Integrated-gut-liver-on-a-chip platform as an in vitro human model of non-alcoholic fatty liver disease.

Non-alcoholic fatty liver disease (NAFLD) afflicts a significant percentage of the population; however, no effective treatments have yet been established because of the unsuitability of in vitro assays and animal experimental models. Here, we present an integrated-gut-liver-on-a-chip (iGLC) platform as an in vitro human model of the gut-liver axis (GLA) by co-culturing human gut and liver cell lines interconnected via microfluidics in a closed circulation loop, for the initiation and progression of NAFLD by treatment with free fatty acids (FFAs) for 1 and 7 days, respectively. Co-cultured Caco-2 gut-mimicking cells and HepG2 hepatocyte-like cells demonstrate the protective effects from apoptosis against FFAs treatment, whereas mono-cultured cells exhibit induced apoptosis. Phenotype and gene expression analyses reveal that the FFAs-treated gut and liver cells accumulated intracellular lipid droplets and show an increase in gene expression associated with a cellular response to copper ions and endoplasmic reticulum stress. As an in vitro human GLA model, the iGLC platform may serve as an alternative to animal experiments for investigating the mechanisms of NAFLD.

Pharmacist perceptions of a "good death" and differences in perception between patients with cancer, oncologists, and oncology nurses: a questionnaire survey.

BACKGROUND: For pharmacists expected to encounter the deaths of many of their patients in the near future, it is important to understand the perception of a "good death" for patients with cancer who are likely to be aware of the circumstances of their poor prognosis. In this study, we clarified pharmacists' perceptions of a "good death" and considered the differences in perception among patients with cancer, oncologists, and oncology nurses. METHODS: From April to June 2022, an anonymous questionnaire survey was conducted on pharmacists working in hospitals and pharmacies and on members of the Japanese Society for Pharmaceutical Palliative Care and Sciences. The questionnaire consisted of 57 questions, called attributes, developed by Miyashita et al. to investigate the perception of "good death" in Japanese cancer medicine. The importance of those attributes was investigated using a 7-point Likert scale. RESULTS: Three thousand four hundred thirty-two pharmacists were made aware of this survey, and 207 participated in the survey. The responses of pharmacists to the 57 questions were very similar to those of the oncologists. Among them, "Fighting against disease until one's last moment" and "Not making trouble for others" had very low importance, which was the most significantly different from the responses of patients with cancer. "Fighting against disease until one's last moment" tended to be significantly underestimated by pharmacists engaged in patient guidance and interview compared to that by pharmacists not engaged in the duty (p = 0.02). Also, when we compared pharmacists with or without qualifications related to cancer and palliative care, there was no significant difference in the importance of "Fighting against disease until one's last moment." However, the importance of "Not making trouble for others" for qualified pharmacists was significantly underestimated (p = 0.04). CONCLUSION: Since pharmacists understand the limits of chemotherapy, they may want to be close to the patient but may not strongly agree with the "Fighting against cancer" component that patients with cancer prefer. It may be necessary to reconsider better ways of approaching the wishes and satisfaction of patients with cancer under the care of medical professionals in the field of oncology.

A pilot study assessing sphingolipids and glycolipids dysmetabolism in idiopathic normal pressure hydrocephalus.

Idiopathic normal pressure hydrocephalus usually exhibits triad of symptoms including gait disturbance, urinary incontinence, and dementia with ventriculomegaly. Currently, its pathogenesis remains to be fully elucidated. To provide a better understanding of this order, we examined whether dysmetabolism of sphingolipids as major lipid components in the brain present in cerebrospinal fluid (CSF) of the patients. Here, we measured various sphingolipidsincluding ceramide and sphingomyelin and glycolipids by electrospray ionization-tandem mass spectrometry in the cerebrospinal fluid of 19 consecutive idiopathic normal pressure hydrocephalus patients, 49 Parkinson's disease patients, and 17 neurologically normal controls. The data showed that there was a significant and specific reduction of all galactosylceramide subspecies levels in idiopathic normal pressure hydrocephalus patients compared with other groups, whereas ceramide and sphingomyelin levels as well as other neutral glycolipids such as glucosylceramide and lactosylceramide were similar in both disease states. Multiple regression analysis of sex and age did not show any correlation with galactosylceramide levels. We also examined whether MMSE scores are correlated with sphingolipid levels in iNPH patients. A specific subspecies of sphingomyelin (d18:1/18:0) only exhibited a statistically significant negative correlation (p = 0.0473, R = -0.4604) with MMSE scores but no other sphingolipids in iNPH patients. These data strongly suggest that myelin-rich galactosylceramide metabolism is severely impaired in idiopathic normal pressure hydrocephalus patients and might serve as the basis of biomarker for this disorder.

Clinical predictors of cardiac syncope in patients with unexplained syncope after the implantation of an insertable cardiac monitor.

Syncope prognosis is related to both its etiology and comorbidities, with cardiac syncope (CS) having higher risks for mortality and cardiovascular events than syncope of non-cardiac causes. Although a novel insertable cardiac monitor (ICM) is an effective diagnostic tool for unexplained syncope, decision regarding ICM implantation with a high pre-test likelihood of CS should contribute to economic cost reduction and avoidance of unnecessary complications. This study aimed to investigate clinical factors associated with CS after ICM implantation in patients with unexplained syncope. This retrospective observational study included 31 consecutive patients with ICM implantation for syncope between September 2016 and August 2021. The initial examinations for syncope included a detailed history, physical examination, blood tests, 12-lead electrocardiograms, and transthoracic echocardiography. Of the 31 patients, 13 (41.9%) experienced recurrent CS during follow-up (676 ± 469 days). Among several clinical factors, syncope-related minor injuries (p = 0.017) and higher brain natriuretic peptide (BNP; p = 0.043) levels were significantly associated with CS. Moreover, multivariable analysis showed that both syncope-related minor injuries (odds ratio, 11.2; 95% confidence interval, 1.4-88.4; p = 0.022) and BNP higher than 64.0 pg/mL (odds ratio, 7.0; 95% confidence interval, 1.1-44.2; p = 0.038) were independent predictors of CS after ICM implantation. In conclusion, a history of minor injury secondary to syncope and higher BNP levels were independent CS predictors in patients receiving ICM for syncope. These results emphasized the utility of ICM implantation early in the diagnostic journey of patients presenting with CS predictors requiring specific treatments.

Multiple Epstein-Barr Virus-associated Gastric Cancers Arising in a Patient with Autoimmune Gastritis.

Epstein-Barr virus-associated gastric cancer (EBVaGC) has been reported to be associated with chronic inflammation of the gastric epithelium caused by Helicobacter pylori infection. Autoimmune gastritis (AIG) is also believed to increase the risk of carcinogenesis. We herein report a case of multiple EBVaGCs that arose in a patient with AIG, highlighting the potential for multiplicity of this entity. In this case, a total of four metachronous EBVaGCs were found after initial Endoscopic submucosal dissection for EBVaGC, all of which were treated endoscopically. This case demonstrates that patients with AIG should be monitored closely for development of EBVaGC.

Phosphoregulation of the transcription factor Mxr1 plays a crucial role in the concentration-regulated methanol induction in Komagataella phaffii.

Methylotrophic yeasts can utilize methanol as the sole carbon and energy source, and the expression of their methanol-induced genes is regulated based on the environmental methanol concentration. Our understanding of the function of transcription factors and Wsc family of proteins in methanol-induced gene expression and methanol sensing is expanding, but the methanol signal transduction mechanism remains undetermined. Our study has revealed that the transcription factor KpMxr1 is involved in the concentration-regulated methanol induction (CRMI) in Komagataella phaffii (Pichia pastoris) and that the phosphorylation state of KpMxr1 changes based on methanol concentration. We identified the functional regions of KpMxr1 and determined its multiple phosphorylation sites. Non-phosphorylatable substitution mutations of these newly identified phosphorylated threonine and serine residues resulted in significant defects in CRMI. We revealed that KpMxr1 receives the methanol signal from Wsc family proteins via KpPkc1 independent of the mitogen-activated protein kinase (MAPK) cascade and speculate that the activity of KpPkc1 influences KpMxr1 phosphorylation state. We propose that the CRMI pathway from Wsc to KpMxr1 diverges from KpPkc1 and that phosphoregulation of KpMxr1 plays a crucial role in CRMI.

Cervical MUC5B and MUC5AC are Barriers to Ascending Pathogens During Pregnancy.

CONTEXT: Cervical excision is a risk factor for preterm birth. This suggests that the cervix plays an essential role in the maintenance of pregnancy. OBJECTIVE: We investigated the role of the cervix through proteomic analysis of cervicovaginal fluid (CVF) from pregnant women after trachelectomy surgery, the natural model of a lack of cervix. METHODS: The proteome compositions of CVF in pregnant women after trachelectomy were compared with those in control pregnant women by liquid chromatography-tandem mass spectrometry and label-free relative quantification. MUC5B/AC expression in the human and murine cervices was analyzed by immunohistochemistry. Regulation of MUC5B/AC expression by sex steroids was assessed in primary human cervical epithelial cells. In a pregnant mouse model of ascending infection, Escherichia coli or phosphate-buffered saline was inoculated into the vagina at 16.5 dpc, and the cervices were collected at 17.5 dpc. RESULTS: The expression of MUC5B/5AC in cervicovaginal fluid was decreased in pregnant women after trachelectomy concomitant with the anatomical loss of cervical glands. Post-trachelectomy women delivered at term when MUC5B/AC abundance was greater than the mean normalized abundance of the control. MUC5B levels in the cervix were increased during pregnancy in both humans and mice. MUC5B mRNA was increased by addition of estradiol in human cervical epithelial cells, whereas MUC5AC was not. In a pregnant mouse model of ascending infection, E. coli was trapped in the MUC5B/AC-expressing mucin of the cervix, and neutrophils were colocalized there. CONCLUSION: Endocervical MUC5B and MUC5AC may be barriers to ascending pathogens during pregnancy.