Germline stem cell (GSC) self-renewal and differentiation into gametes is regulated by both intrinsic factors in the germ line as well as extrinsic factors from the surrounding somatic niche. dWnt4, in the escort cells of the adult somatic niche promotes GSC differentiation using the canonical β-catenin-dependent transcriptional pathway to regulate escort cell survival, adhesion to the germ line and downregulation of self-renewal signaling. Here, we show that in addition to the β-catenin-dependent canonical pathway, dWnt4 also uses downstream components of the Wnt non-canonical pathway to promote escort cell function earlier in development. We find that the downstream non-canonical components, RhoA, Rac1 and cdc42, are expressed at high levels and are active in escort cell precursors of the female larval gonad compared to the adult somatic niche. Consistent with this expression pattern, we find that the non-canonical pathway components function in the larval stages but not in adults to regulate GSC differentiation. In the larval gonad, dWnt4, RhoA, Rac1 and cdc42 are required to promote intermingling of escort cell precursors, a function that then promotes proper escort cell function in the adults. We find that dWnt4 acts by modulating the activity of RhoA, Rac1 and cdc42, but not their protein levels. Together, our results indicate that at different points of development, dWnt4 switches from using the non-canonical pathway components to using a β-catenin-dependent canonical pathway in the escort cells to facilitate the proper differentiation of GSCs.
Introduction: Alpha-fetoprotein (AFP) functions in utero to inhibit estrogen-mediated growth in fetal tissue. AFPep is a 9-amino acid cyclized form of the active site in AFP that impedes phosphorylation and activation of ERα. Recent studies demonstrate that AFPep inhibits estrogen-dependent growth in MCF-7 and T47D xenografts and decreases mammary tumor burden in estrogen-exposed ACI rats. It is important to identify biomarkers that predict the efficacy of AFPep. Dimerized, phosphorylated ERα (pERα) transcribes TRIM25, which is associated with breast cancer metastasis and decreased survival. KLF5 is a transcription factor downstream of pERα and is associated with breast and cervical cancer proliferation, invasiveness, and migration. We hypothesize that treatment with AFPep inhibits expression of TRIM25 and KLF5 and that these are predictive biomarkers for AFPep-mediated inhibition of estrogen-dependent growth. Methods: Expression of ERα, pERα, TRIM25, and KLF5 were assessed in three estrogen-sensitive models: immature mouse uterus, MCF7 xenografts, and canine breast tumors. Immature mice were treated with saline only, AFPep (100 μg) only, estrogen, or estrogen plus AFPep (100 μg). Twenty-four hours post treatment, uteri were removed, weighed, and blotted for biomarkers. Mice bearing MCF7 xenografts were treated with estrogen or estrogen with AFPep and biopsied after 14 days. Finally, two dogs, one with a simple tubular mammary adenoma and the other with a grade II mixed mammary carcinoma, were treated with AFPep for seven days. Results: Immature mice treated with estrogen had greater uterine-to-body weight ratio, increased ratio of pERα to total ERα, and increased KLF5 and TRIM25 expression compared to mice treated with saline alone or AFPep alone. Uteri from mice treated with estrogen and AFPep showed decreased uterine-to-body weight ratio, decreased ratio of pERα to total ERα, and decreased KLF5 and TRIM25 expression compared to uteri treated with estrogen alone. MCF7 xenografts from mice treated with estrogen and AFPep showed decreased ratio of pERα to total ERα compared to mice treated with estrogen alone. AFPep treatment of xenograft-bearing mice decreased expression of TRIM25 and KLF5 in the tumor. In the clinical canine mammary tumor study, a mixed carcinoma expressed pERα and treatment of that dog with AFPep decreased the expression of both KLF5 and TRIM25 in that tumor. A dog with a simple tubular adenoma did not express pERα, and treatment of that dog with AFPep had little impact on the expression of TRIM25 and KLF5 in the tumor. Conclusion: We conclude that AFPep inhibits estrogen-mediated growth and the ratio of pERα to total ERα. KLF5 and TRIM25 also serve as predictive markers for the efficacy of AFPep in uterus and breast tissue respectively. Citation Format: Kanthi J. Bommareddy, Anusri Kadakuntla, Michael Kuna, Priya Shivraj, Roman Ginnan, Ann Hohenhaus, James Bennett, Thomas Andersen. Trim25 and KLF5 expression predict AFPep-mediated inhibition of estrogen-dependent growth [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2968.
INTRODUCTION: We present cases of esophageal motility disorders associated with sleeve gastrectomy and identify the role of high-resolution manometry (HRM) in perioperative diagnosis and management. CASE DESCRIPTION/METHODS: Case 1: A 44-year old female with history of morbid obesity and gastric reflux presented with persistent symptoms of regurgitation, vomiting, and refractory reflux after undergoing a sleeve gastrectomy two years prior. Upper endoscopy showed slight resistance to endoscope passage at the lower esophageal sphincter and HRM evaluation was consistent with Type II achalasia, a finding confirmed by barium esophagram. The patient underwent a Heller myotomy with conversion into gastric bypass. On follow-up, patient noted no further recurrence of her presenting symptoms. Case 2: 52-year old female with multiple comorbidities presented with refractory dysphagia. She had undergone sleeve gastrectomy 5 years prior and subsequently developed worsening postprandial chest pain and solid food dysphagia. Outside work up of her dysphagia including multiple endoscopies and an esophagram were negative. On present evaluation, a 13 mm Barium pill traversed the esophagus without delay, and HRM showed a hypercontractile esophagus with high normal integrated relaxation pressure. The patient was started on antispasmodic therapy with symptom improvement. DISCUSSION: Laparoscopic sleeve gastrectomy is a bariatric surgical procedure for rapid weight loss in those classified as morbidly obese. The procedure leaves patients with a narrow tubular stomach roughly the diameter of the esophagus. Post operatively, patients have been noted to experience dysphagia, odynophagia, and most commonly (19.7% of cases) reflux. In our cases, patients experienced severe esophageal dysmotility which may have been caused by post-operative alterations of gastric anatomy and pressures. The mechanism behind post-operative motility disorders is unclear, and some studies have suggested that the procedure may unmask previously latent or subclinical disorders rather than induce them. We believe that HRM should be used as part of high value care in evaluating gastric sleeve patients with esophageal symptoms, especially before surgical revisions. Braghetto, I., et al. OBES SURG (2010) 20: 357. https://doi-org.elibrary.amc.edu/10.1007/s11695-009-0040-3.
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