Publications by authors named "Sotirios G Doukas"

24 Publications

  • Page 1 of 1

Rapidly Progressing Primary Membranous Nephropathy in a Hispanic Male With Elevated Levels of Anti-phospholipase A2 Receptor Antibodies.

Cureus 2021 Jun 11;13(6):e15594. Epub 2021 Jun 11.

Internal Medicine, Saint Peter's University Hospital, New Brunswick, USA.

A 49-year-old Hispanic male presented to the Emergency Department with progressively worsening swelling in his extremities for the past two months. Physical examination was significant for diffuse edema, with concomitant initial laboratory tests revealing hypoalbuminemia, hypercholesteremia, and proteinuria. A renal biopsy was performed, and the histopathology confirmed a diagnosis of membranous nephropathy through immunofluorescence and electron microscopy. Anti-phospholipase A2 receptor (anti-PLA2R) antibodies were detected on immunofluorescence, as well as high levels being discovered in the patient's serum, indicating a diagnosis of primary membranous nephropathy. The patient underwent adequate diuresis and was discharged. The patient presented six months later due to severe anasarca with laboratory tests indicating a rapid decline in renal function. He was then started on immunosuppressive therapy. Our rare case of a Hispanic male presenting with rapidly deteriorating renal function secondary to primary membranous nephropathy seeks to highlight the possibility of using anti-PLA2R antibodies as a marker for early initiation of immunosuppressive therapy as well as to encourage additional research on the course of disease progression in the Hispanic population.
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http://dx.doi.org/10.7759/cureus.15594DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8272928PMC
June 2021

Controlled ovarian stimulation therapy as a potential risk for the development and progression of renal cell carcinomas: A case report and literature review.

Mol Clin Oncol 2021 Jul 23;15(1):140. Epub 2021 May 23.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT 06510, USA.

Renal Cell Carcinoma (RCC) is the most common type of cancer in the kidney and is mostly asymptomatic. Previous studies have supported the important role of sex hormones in RCC pathophysiology and that targeted hormone receptor therapy, such as estrogen receptor targeting, is a promising treatment strategy. However, to the best of our knowledge, it remains unknown whether hormonal therapy, such as controlled ovarian stimulation for fertilization, serves a role in the development and progression of RCC. The present report describes a case of RCC developed after a fertility stimulation therapy and provides a summary of the known literature on the role of hormone receptors in the development and progression of RCC. A 35-year-old woman received fertility stimulation treatment with follitropin alfa 900 units, human chorionic gonadotropic hormone 5,000 units, injectable leuprolide 1 mg/0.2 ml and cetrotide 0.25 mg. The patient presented to the hospital with shortness of breath and weight loss. The patient had no known genetic predisposition or family history of malignancies and no exposure to chemicals. The patient never used tobacco, alcohol or recreational drugs. Imaging revealed a 17x19 mm, heterogeneously enhancing, and partially exophytic mass in the right kidney. After partial nephrectomy, the pathological evaluation confirmed the diagnosis of clear cell RCC. To the best of our knowledge, this was the first time that a case of ovarian stimulation therapy was associated with the development of RCC. This case raises concerns about the potential oncogenic effect of controlled ovarian stimulation therapy in RCC promotion, suggesting a need for systematic research to clarify the clinical significance of existing pre-clinical data.
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http://dx.doi.org/10.3892/mco.2021.2302DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8165689PMC
July 2021

Psoas Abscess Presented as Right Hip Pain in a Young Adult With Crohn's Disease.

Cureus 2021 Feb 5;13(2):e13162. Epub 2021 Feb 5.

Department of Medicine, Saint Peter's University Hospital, New Brunswick, USA.

A 27-year-old man with a past medical history of Crohn's disease presented in the Emergency Department complaining of right hip pain that has been going on for one month. At presentation, the patient was tachycardic. Physical examination revealed a positive psoas sign. Laboratory tests showed elevated white blood cells, C-reactive protein, and erythrocyte sedimentation rate. Computed tomography of the abdomen and pelvis revealed ileo-psoas fistula and psoas abscess. This rare case aims to provide awareness that intra-abdominal pathology should always be suspected in patients with referred hip pain and Crohn's disease. A thorough physical examination including maneuvers for assessment of possible iliopsoas inflammation should be effectively performed at the bedside to determine the likelihood of the condition and proper imaging should follow to confirm the diagnosis.
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http://dx.doi.org/10.7759/cureus.13162DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7935276PMC
February 2021

Weakly Acidic Bile Is a Risk Factor for Hypopharyngeal Carcinogenesis Evidenced by DNA Damage, Antiapoptotic Function, and Premalignant Dysplastic Lesions In Vivo.

Cancers (Basel) 2021 Feb 18;13(4). Epub 2021 Feb 18.

The Yale Larynx Laboratory, Department of Surgery (Otolaryngology), Yale School of Medicine, New Haven, CT 06510, USA.

Background: There is recent in vivo discovery documenting the carcinogenic effect of bile at strongly acidic pH 3.0 in hypopharynx, while in vitro data demonstrate that weakly acidic bile (pH 5.5) has a similar oncogenic effect. Because esophageal refluxate often occurs at pH > 4.0, here we aim to determine whether weakly acidic bile is also carcinogenic in vivo.

Methods: Using 32 wild-type mice C57B16J, we performed topical application of conjugated primary bile acids with or without unconjugated secondary bile acid, deoxycholic acid (DCA), at pH 5.5 and controls, to hypopharyngeal mucosa (HM) twice per day, for 15 weeks.

Results: Chronic exposure of HM to weakly acidic bile, promotes premalignant lesions with microinvasion, preceded by significant DNA/RNA oxidative damage, γH2AX (double strand breaks), NF-B and p53 expression, overexpression of -2, and elevated and mRNAs, compared to controls. Weakly acidic bile, without DCA, upregulates the "oncomirs", -21 and -155. The presence of DCA promotes , , and overexpression, and a significant downregulation of "tumor suppressor" -451a.

Conclusion: Weakly acidic pH increases the risk of bile-related hypopharyngeal neoplasia. The oncogenic properties of biliary esophageal reflux on the epithelium of the upper aerodigestive tract may not be fully modified when antacid therapy is applied. We believe that due to bile content, alternative therapeutic strategies using specific inhibitors of relevant molecular pathways or receptors may be considered in patients with refractory GERD.
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http://dx.doi.org/10.3390/cancers13040852DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7923205PMC
February 2021

Achalasia in Myotonic Dystrophy.

Am J Gastroenterol 2021 02;116(2):429-431

Division of Gastroenterology, Rutgers/Saint Peter's University Hospital, New Brunswick, New Jersey, USA.

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http://dx.doi.org/10.14309/ajg.0000000000000951DOI Listing
February 2021

E-cigarette or vaping induced lung injury: A case series and literature review.

Toxicol Rep 2020 3;7:1381-1386. Epub 2020 Oct 3.

Department of Internal Medicine, Saint Peter's University Hospital, 254 Easton Ave, New Brunswick, NJ, 08901, USA.

Introduction: Recently, a rapidly increasing number of e-cigarette or vaping induced lung injury (EVALI) has been reported across the nation. Given the ongoing epidemic, it has been suggested that specific chemical substances used as additives in e-cigarettes could be highly related to EVALI. A history of vaping with positive radiographic changes and low suspicion for active infection are requirements for diagnosis but it still remains a diagnosis of exclusion. The course of the disease, mechanism of lung injury and the optimal management options need to be better understood. Here we aimed to discuss the clinical characteristics recognized in a case series of ten hospitalized EVALI patients with radiological findings of lung injury and provide an up today summary of the known literature of EVALI-induced lung injury.

Methods: A retrospective chart review was conducted on ten patients who presented to Saint Peter's University Hospital in New Brunswick, NJ from July 2019 to February 2020, with a mean hospital stay of five days. According to the CDC recommended definition of the disease, our cases met the current working definition of confirmed or probable cases of EVALI.

Results: Ten patients, with mean age 30.8 years (50 % male) and average years of vaping 1.708 with 60 % endorsing a simultaneous history of cannabis-related products use, went under a retrospective review. 3/10 (30 %) had documented medically-managed pulmonary disease history, 8/10 (80 %) presented with the respiratory-related chief complaint, 6/10 (60 %) presented with gastrointestinal symptoms and 7/10 (70 %) had constitutional symptoms. All patients (100 %) were found to have bilateral ground-glass opacities on chest imaging. 9/10 were admitted, 6/10 (60 %) had an oxygen saturation of <95 % requiring oxygen supplementation with 4/10 managed in the intensive care unit.

Conclusion: EVALI patients with radiological findings of lung injury, although mainly present respiratory symptoms, may very often appear with constitutional and gastrointestinal symptoms. Based on the existing literature and our data it is argued that EVALI may be misdiagnosed and that closer monitoring is required to determine optimal diagnostic and therapeutic management of this condition. Our data and the existing literature suggest that laboratory and epidemiologic findings can be contributory for the diagnosis of the disease.
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http://dx.doi.org/10.1016/j.toxrep.2020.09.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7577885PMC
October 2020

SARS-CoV-2 pathophysiology and its clinical implications: An integrative overview of the pharmacotherapeutic management of COVID-19.

Food Chem Toxicol 2020 Dec 30;146:111769. Epub 2020 Sep 30.

Department of Toxicology, University of Medicine and Pharmacy of Craiova, 200349, Craiova, Romania. Electronic address:

Common manifestations of COVID-19 are respiratory and can extend from mild symptoms to severe acute respiratory distress. The severity of the illness can also extend from mild disease to life-threatening acute respiratory distress syndrome (ARDS). SARS-CoV-2 infection can also affect the gastrointestinal tract, liver and pancreatic functions, leading to gastrointestinal symptoms. Moreover, SARS-CoV-2 can cause central and peripheral neurological manifestations, affect the cardiovascular system and promote renal dysfunction. Epidemiological data have indicated that cancer patients are at a higher risk of contracting the SARS-CoV-2 virus. Considering the multitude of clinical symptoms of COVID-19, the objective of the present review was to summarize their pathophysiology in previously healthy patients, as well as in those with comorbidities. The present review summarizes the current, though admittedly fluid knowledge on the pathophysiology and symptoms of COVID-19 infection. Although unclear issues still remain, the present study contributes to a more complete understanding of the disease, and may drive the direction of new research. The recognition of the severity of the clinical symptoms of COVID-19 is crucial for the specific therapeutic management of affected patients.
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http://dx.doi.org/10.1016/j.fct.2020.111769DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7833750PMC
December 2020

The temporal effects of topical NF- inhibition, in the prevention of bile-related oncogenic mRNA and miRNA phenotypes in murine hypopharyngeal mucosa: a preclinical model.

Oncotarget 2020 Sep 1;11(35):3303-3314. Epub 2020 Sep 1.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT, USA.

Supraesophageal bile reflux at strongly acidic pH can cause hypopharyngeal squamous cell cancer, through activation of the oncogenic NF-κB-related pathway. We hypothesize that topical pre- or post-application of pharmacologic NF-κB inhibitor, BAY 11-7082 (0.25 μmol), on murine (C57BL/6J) HM (twice a day for 10 days) can effectively inhibit acidic bile (10 mmol/l; pH 3.0) induced oncogenic molecular events, similar to prior findings. We demonstrate that the administration of BAY 11-7082, either before or after acidic bile, eliminates NF-κB activation, prevents overexpression of and deregulations of , , linked to bile reflux-related hypopharyngeal cancer. Pre- but not post-application of NF-κB inhibitor, significantly blocks overexpression of and prostaglandin H synthases 2 (), and reverses phenotypes, supporting its early bile-induced pro-inflammatory effect. We thus provide novel evidence that topical administration of a pharmacological NF-κB inhibitor, either before or after acidic bile exposure can successfully prevent its oncogenic mRNA and miRNA phenotypes in HM, supporting the observation that co-administration of NF-κB inhibitor may not be essential in preventing early bile-related oncogenic events and encouraging a capacity for further translational exploration.
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http://dx.doi.org/10.18632/oncotarget.27706DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7476734PMC
September 2020

The in vivo preventive and therapeutic properties of curcumin in bile reflux-related oncogenesis of the hypopharynx.

J Cell Mol Med 2020 09 21;24(18):10311-10321. Epub 2020 Jul 21.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT, USA.

Bile at strongly acidic pH exerts a carcinogenic effect on the hypopharynx, based upon recent pre-clinical studies that support its role as an independent risk factor. We recently demonstrated in vitro that curcumin can prevent oncogenic profile of bile in human hypopharyngeal cells, by inhibiting NF-κB. We hypothesize that topically applied curcumin to the hypopharynx can similarly block early oncogenic molecular events of bile, by inhibiting NF-κB and consequently altering the expression of genes with oncogenic function. Using Mus musculus (C57Bl/6J), we topically applied curcumin (250 μmol/L; three times per day; 10 days) to the hypopharynx, 15 minutes before, 15 minutes after or in combination with bile acids (pH 3.0). Immunohistochemical analysis and qPCR revealed that topically applied curcumin either before, after or in combination with acidic bile exposure significantly suppressed its induced NF-κB activation in regenerating epithelial cells, and overexpression of Rela, Bcl2, Egfr, Stat3, Wnt5a, Tnf, Il6, Ptgs2. Akt1 was particularly inhibited by curcumin when applied simultaneously with bile. We provide novel evidence into the preventive and therapeutic properties of topically applied curcumin in acidic bile-induced early oncogenic molecular events in hypopharyngeal mucosa, by inhibiting NF-κB, and shaping future translational development of effective targeted therapies using topical non-pharmacologic inhibitors of NF-κB.
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http://dx.doi.org/10.1111/jcmm.15640DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7521262PMC
September 2020

The Progressive Mutagenic Effects of Acidic Bile Refluxate in Hypopharyngeal Squamous Cell Carcinogenesis: New Insights.

Cancers (Basel) 2020 Apr 25;12(5). Epub 2020 Apr 25.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, 310 Cedar Street (BML212), New Haven, CT 06510, USA.

Cancers of the laryngopharynx represent the most devastating of the head and neck malignancies and additional risk factors are now epidemiologically linked to this disease. Using an in vivo model ( C57Bl/6J), we provide novel evidence that acidic bile (pH 3.0) progressively promotes invasive cancer in the hypopharynx. Malignant lesions are characterized by increasing: i) oxidative DNA-damage, ii) γH2AX expression, iii) NF-κB activation, and iv) p53 expression. Histopathological changes observed in murine hypopharyngeal mucosa exposed to acidic bile were preceded by the overexpression of , , , , , , and the deregulation of , , , , , . This is the first study to document that acidic bile is carcinogenic in the upper aerodigestive tract. We showed that oxidative DNA-damage produced by acidic bile in combination with NF-κB-related anti-apoptotic deregulation further supports the underlying two-hit hypothesized mechanism. Just as importantly, we reproduced the role of several biomarkers of progression that served as valuable indicators of early neoplasia in our experimental model. These findings provide a sound basis for proposing translational studies in humans by exposing new opportunities for early detection and prevention.
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http://dx.doi.org/10.3390/cancers12051064DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7281001PMC
April 2020

The Effect of NNK, A Tobacco Smoke Carcinogen, on the miRNA and Mismatch DNA Repair Expression Profiles in Lung and Head and Neck Squamous Cancer Cells.

Cells 2020 04 21;9(4). Epub 2020 Apr 21.

Department of Forensic Sciences and Laboratory of Toxicology, Medical School, University of Crete, 71003 Heraklion, Greece.

Tobacco smoking is a common risk factor for lung cancer and head and neck cancer. Molecular changes such as deregulation of miRNA expression have been linked to tobacco smoking in both types of cancer. Dysfunction of the Mismatch DNA repair (MMR) mechanism has also been associated with a poor prognosis of these cancers, while a cross-talk between specific miRNAs and MMR genes has been previously proposed. We hypothesized that exposure of lung and head and neck squamous cancer cells (NCI and FaDu, respectively) to tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is capable of altering the expression of MSH2 and MLH1, key MMR components, by promoting specific miRNA deregulation. We found that either a low (1 μM) or high (2 μM) dose of NNK induced significant upregulation of "oncomirs" miR-21 and miR-155 and downregulation of "tumor suppressor" miR-422a, as well as the reduction of MMR protein and mRNA expression, in NCI and FaDu, compared to controls. Inhibition of miR-21 restored the NNK-induced reduced MSH2 phenotype in both NCI and FaDu, indicating that miR-21 might contribute to MSH2 regulation. Finally, NNK exposure increased NCI and FaDu survival, promoting cancer cell progression. We provide novel findings that deregulated miR-21, miR-155, and miR-422a and MMR gene expression patterns may be valuable biomarkers for lung and head and neck squamous cell cancer progression in smokers.
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http://dx.doi.org/10.3390/cells9041031DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7226174PMC
April 2020

The role of bile reflux and its related NF-κB activated pathway in progression of hypopharyngeal squamous cell cancer.

Oral Oncol 2020 06 2;105:104668. Epub 2020 Apr 2.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT, USA. Electronic address:

Background: Prognosis for hypopharyngeal cancer is usually poor, and recurrence is common. Identifying new factors or related mechanisms that promote its progression may have clinical implications. Although, recent studies support bile reflux in hypopharyngeal carcinogenesis, it remains to be explored how bile and its related NF-κB activated pathway may further affects its progression in already established hypopharyngeal cancer.

Methods: Hypopharyngeal squamous cell carcinoma (HSCC) cell lines, FaDu and UMSCC11A, both negative for HPV, were repetitively exposed to bile acids (400 μM) at variable pH points (4.0, 5.5 and 7.0). Immunofluorescence, western blotting, luciferase assay, and qPCR were used to detect NF-κB activation, bcl-2 overexpression and gene expression.

Results: Bile at strongly acidic pH (4.0) potentiated the activation of NF-κB and its related mRNA phenotype in HSCC cells. IL-6, TNF-α, and BCL2 were found among the highest overexpressed genes as was previously found in HSCCs excised from patients with documented biliary reflux. An enhanced transcriptional activity of EGFR, RELA, STAT3, and WNT5Α and higher survival rates were observed in HSCC cells exposed to acidic bile compared to those exposed to bile at weakly acidic or neutral pH.

Conclusion: Our novel findings support the observation that bile reflux has the potential for actively influencing the progression of hypopharyngeal cancer, mediated by NF-κB. In patients with hypopharyngeal cancer and known gastroesophageal reflux disease, antacid therapy may exert a role in furthering control of disease recurrence and progression.
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http://dx.doi.org/10.1016/j.oraloncology.2020.104668DOI Listing
June 2020

Rare amyloidoma of the tongue base: A case report and review of the literature.

Mol Clin Oncol 2020 Mar 9;12(3):258-262. Epub 2020 Jan 9.

Department of Otorhinolaryngology, University of Medicine and Pharmacy of Craiova, 200349 Craiova, Romania.

Localized amyloidosis is a rare condition characterized by the deposition of misfolding protein in a tissue, without other systemic manifestations. Only a small number of cases of localized amyloidosis of the tongue have been reported to date, in contrast to systemic amyloidosis, in which localization on the tongue is common. This study presents a rare case of localized amyloidosis of the tongue (amyloidoma) and provides a summary of the known literature of localized amyloidosis. This study describes the case of a 36-year-old female who presented with a swelling of the tongue base. The diagnosis of amyloidoma was made based on the findings of the physical examination, head and neck MRI findings and the histopathological examination with Congo red stain under polarized light. The histopathological diagnosis was as follows: Localized lambda light-chain amyloidosis. A thorough physical examination was performed by the ENT and Hematology/Oncology departments, without revealing signs of systemic disease. A series of hematological and imaging tests were also performed to verify that there was no sign of systemic involvement. The patient declined surgical excision and the 2-year follow-up did not reveal any changes in tumor dimension. Although the etiology of localized amyloidosis is yet not clear, the prolonged reaction of tissue plasma cells to environmental antigens may be a causative factor for the initiation of the neoplastic process.
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http://dx.doi.org/10.3892/mco.2020.1972DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7016517PMC
March 2020

Biliary reflux as a causal factor in hypopharyngeal carcinoma: New clinical evidence and implications.

Cancer 2019 10 16;125(20):3554-3565. Epub 2019 Jul 16.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, Connecticut.

Background: Recent preclinical explorations strongly support the tumorigenic potential of bile on laryngopharyngeal mucosa. Herein, the authors describe, in bile-related human hypopharyngeal squamous cell carcinoma (HSCC), NF-κB-related messenger RNA (mRNA) and microRNA (miRNA) oncogenic phenotypes similar to those previously identified in acidic bile-exposed premalignant murine hypopharyngeal mucosa.

Methods: In this pilot study, the authors included human HSCC specimens paired with their adjacent normal tissue (ANT) derived from 3 representative patients with documented biliary laryngopharyngeal reflux (bile[+]) compared with 5 control patients without signs of bile reflux disease (bile[-]). Immunohistochemical, quantitative polymerase chain reaction, and miRNA analyses were used to detect the levels of activated NF-κB and expression levels of STAT3, EGFR, BCL2, WNT5A, IL-6, IL-1B, ΔNp63, cREL, TNF-α, TP53, NOTCH1, NOTCH2, NOTCH3, miR-21, miR-155, miR-192, miR-34a, miR-375, miR-451a, miR-489, miR-504, and miR-99a.

Results: Bile(+) HSCC demonstrated an intense NF-κB activation accompanied by significant overexpression of RELA(p65), EGFR, STAT3, BCL-2, cREL, ΔNp63, WNT5A, IL-6, and IL1B; upregulation of oncomir miR-21; and downregulation of tumor suppressor miR-375 compared with their respective ANTs. Bile(+) HSCC demonstrated significantly higher mRNA levels of all the analyzed genes, particularly RELA(p65), IL-6, EGFR, and TNF-α compared with bile(-) tumors. The miR-21/miR-375 ratio, which previously has been linked to tumor aggressiveness, was found to be >260-fold and >30-fold higher, respectively, in bile(+) HSCCs compared with their ANTs and bile(-) tumors.

Conclusions: Although limitations apply to this pilot study due to the small number of patients with HSCC, the novel findings suggest that a history of bile as a component of esophageal reflux disease may represent an independent risk factor for hypopharyngeal carcinogenesis.
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http://dx.doi.org/10.1002/cncr.32369DOI Listing
October 2019

Biliary tumorigenic effect on hypopharyngeal cells is significantly enhanced by pH reduction.

Cancer Med 2019 08 7;8(9):4417-4427. Epub 2019 Jun 7.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, Connecticut, USA.

Biliary reflux has been considered a potential risk factor in upper aerodigestive tract malignancies. It is not yet clearly known how pH affects the bile-induced activation of NF-κB and its related oncogenic pathway previously linked to hypopharyngeal carcinogenesis. In this study, repetitive applications of conjugated primary bile acids with unconjugated secondary bile acid, deoxycholic acid (DCA), on human hypopharyngeal primary cells reveal that strongly acidic pH (4.0) optimally enhances the tumorigenic effect of bile, by inducing activation of NF-κB, STAT3 nuclear translocation, bcl-2 overexpression and significant overexpression of the oncogenic mRNA phenotype, compared to weakly acidic pH (5.5) or neutral pH (7.0). As the pH becomes less acidic the partially activated primary bile acids and activated DCA begin to exert their influence; however, with significantly less intensity compared to bile acids at strongly acidic pH. Our findings suggest that biliary tumorigenic effect is strongly pH dependent. Controlling pH during reflux events may be therapeutically effective in reducing the potential risk of bile-induced hypopharyngeal cancer.
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http://dx.doi.org/10.1002/cam4.2194DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6675744PMC
August 2019

Temporal characteristics of NF-κB inhibition in blocking bile-induced oncogenic molecular events in hypopharyngeal cells.

Oncotarget 2019 May 21;10(36):3339-3351. Epub 2019 May 21.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT, USA.

Biliary esophageal reflux at acidic pH is considered a risk factor in laryngopharyngeal cancer. We previously showed the key role NF-B in mediating acidic bile-induced pre-neoplastic events in hypopharyngeal cells, and that co-administration of specific NF-B inhibitor, BAY 11-7082, together with acidic bile, can effectively prevent its related oncogenic molecular effects. We hypothesize that the addition of BAY 11-7082 (10μM) either before or after application of acidic bile (400μM conjugated bile acids; pH 4.0), is capable of comparably blocking acidic bile-induced oncogenic molecular phenotypes in murine hypopharyngeal primary cells. We performed immunofluorescence, luciferase assay, western blot and qPCR analysis, demonstrating that 15-min of pre- or post-application of BAY 11-7082 effectively inhibits acidic bile-induced NF-B activation, transcriptional activation of W, "upregulation" of "oncomirs" miR-21, miR-155, miR-192 and "downregulation" of "tumor suppressor" miR-34a, miR-375, miR-451a. Our observations support the understanding that acidic bile-induced deregulation of anti-apoptotic or oncogenic factors, , , miR-21, miR-155, miR-375, is highly NF-B-dependent, showing that even post-application of inhibitor can suppress their deregulation. In conclusion, application of specific NF-κB inhibitor, has the capability of adequately blocking the early oncogenic molecular events produced by acidic bile whether it is applied pre or post exposure. In addition to therapeutic implications these findings provide a window of observation into the complex kinetics characterizing the mechanistic link between acidic bile and early neoplasia. Although BAY 11-7082 itself may not be suitable for clinical use, the application of other NF-κB inhibitors merits exploration.
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http://dx.doi.org/10.18632/oncotarget.26917DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6534360PMC
May 2019

In search of a longitudinal animal model of evoked swallow function.

Laryngoscope Investig Otolaryngol 2018 Jun 14;3(3):191-197. Epub 2018 May 14.

Yale University School of Medicine New Haven Connecticut U.S.A.

Background: A malfunction or impairment of swallow function can potentiate aspiration events and interfere with both quality of life and survival. Establishing an animal model for swallow research would provide a better understanding of its pathophysiology and would also allow for the development and validation of physiologically based clinical interventions to improve swallow function. Two requirements define the ideal model for longitudinal exploration: 1) identification of species similar to human in form and function; and 2) provision for reliable and reproducible evoked swallow under general anesthesia and one that would also support a longitudinal study design.

Objective: We hypothesize that an anesthetized porcine model under dexmedetomidine-based or ketamine-based anesthesia will support a reproducible and stable evoked swallow response.

Methods: Seven neutered male Yorkshire pigs were anesthetized using combinations of dexmedetomidine-based or ketamine-based anesthesia for induction and maintenance of anesthesia during the experimental portion of our study. Single stimulation of iSLN or vagus nerve, bilateral simultaneous single stimulation of iSLN or vagus nerve, and stimulus trains applied to afferent nerves were performed.

Results: None of the seven pigs demonstrated evoked swallow events, both during inhalational anesthesia (1.0 MAC) or during post-washout intravenous anesthesia (dexmedetomidine, ketamine/fentanyl or ketamine alone).

Conclusion: Our results support a high degree of organizational neurophysiologic complexity characterizing the swallow reflex and highlight the challenges and limitations of intraoperative study in survival models.

Level Of Evidence: NA , 2018.
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http://dx.doi.org/10.1002/lio2.161DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6057227PMC
June 2018

Curcumin prevents the bile reflux-induced NF-κB-related mRNA oncogenic phenotype, in human hypopharyngeal cells.

J Cell Mol Med 2018 09 17;22(9):4209-4220. Epub 2018 Jun 17.

Department of Surgery, The Yale Larynx Laboratory, Yale School of Medicine, New Haven, CT, USA.

The presence of bile is not an uncommon finding in acidic oesophageal and extra-oesophageal refluxate, possibly affecting the hypopharyngeal mucosa and leading to neoplastic events. We recently demonstrated that acidic bile (pH ≤ 4.0) can induce NF-κB activation and oncogenic mRNA phenotype in normal hypopharyngeal cells and generate premalignant changes in treated hypopharyngeal mucosa. We hypothesize that curcumin, a dietary inhibitor of NF-κB, may effectively inhibit the acidic bile-induced cancer-related mRNA phenotype, in treated human hypopharyngeal primary cells (HHPC), supporting its potential preventive use in vivo. Luciferase assay, immunofluorescence, Western blot, qPCR and PCR microarray analysis were used to explore the effect of curcumin in HHPC exposed to bile (400 μmol/L) at acidic and neutral pH. Curcumin successfully inhibited the acidic bile-induced NF-κB signalling pathway (25% of analysed genes), and overexpression of NF-κB transcriptional factors, c-REL, RELA(p65), anti-apoptotic bcl-2, oncogenic TNF-α, EGFR, STAT3, WNT5A, ΔNp63 and cancer-related IL-6. Curcumin effectively reduced bile-induced bcl-2 overexpression at both acidic and neutral pH. Our novel findings suggest that, similar to pharmacologic NF-κB inhibitor, BAY 11-7082, curcumin can suppress acidic bile-induced oncogenic mRNA phenotype in hypopharyngeal cells, encouraging its future in vivo pre-clinical and clinical explorations in prevention of bile reflux-related pre-neoplastic events mediated by NF-κB.
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http://dx.doi.org/10.1111/jcmm.13701DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6111812PMC
September 2018

In Vivo Short-Term Topical Application of BAY 11-7082 Prevents the Acidic Bile-Induced mRNA and miRNA Oncogenic Phenotypes in Exposed Murine Hypopharyngeal Mucosa.

Neoplasia 2018 04 9;20(4):374-386. Epub 2018 Mar 9.

Department of Surgery, Yale Larynx Laboratory, Yale School of Medicine, New Haven, CT, USA. Electronic address:

Purpose: Bile-containing gastroesophageal reflux may promote cancer at extraesophageal sites. Acidic bile can accelerate NF-κB activation and molecular events, linked to premalignant changes in murine hypopharyngeal mucosa (HM). We hypothesize that short-term in vivo topical application of NF-κB inhibitor BAY 11-7082 can prevent acidic bile-induced early preneoplastic molecular events, suggesting its potential role in disease prevention.

Experimental Design: We topically exposed HM (C57Bl/6j wild-type) to a mixture of bile acids at pH 3.0 with and without BAY 11-7082 3 times/day for 7 days. We used immunofluorescence, Western blotting, immunohistochemistry, quantitative polymerase chain reaction, and polymerase chain reaction microarrays to identify NF-κB activation and its associated oncogenic mRNA and miRNA phenotypes, in murine hypopharyngeal cells in vitro and in murine HM in vivo.

Results: Short-term exposure of HM to acidic bile is a potent stimulus accelerating the expression of NF-κB signaling (70 out of 84 genes) and oncogenic molecules. Topical application of BAY 11-7082 sufficiently blocks the effect of acidic bile. BAY 11-7082 eliminates NF-κB activation in regenerating basal cells of acidic bile-treated HM and prevents overexpression of molecules central to head and neck cancer, including bcl-2, STAT3, EGFR, TNF-α, and WNT5A. NF-κB inhibitor reverses the upregulated "oncomirs" miR-155 and miR-192 and the downregulated "tumor suppressors" miR-451a and miR-375 phenotypes in HM affected by acidic bile.

Conclusion: There is novel evidence that acidic bile-induced NF-κB-related oncogenic mRNA and miRNA phenotypes are generated after short-term 7-day mucosal exposure and that topical mucosal application of BAY 11-7082 can prevent the acidic bile-induced molecular alterations associated with unregulated cell growth and proliferation of hypopharyngeal cells.
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http://dx.doi.org/10.1016/j.neo.2018.02.001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5909679PMC
April 2018

NF-κB inhibition reverses acidic bile-induced miR-21, miR-155, miR-192, miR-34a, miR-375 and miR-451a deregulations in human hypopharyngeal cells.

J Cell Mol Med 2018 05 8;22(5):2922-2934. Epub 2018 Mar 8.

The Yale Larynx laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT, USA.

We previously demonstrated that acidic bile activates NF-κB, deregulating the expression of oncogenic miRNA markers, in pre-malignant murine laryngopharyngeal mucosa. Here, we hypothesize that the in vitro exposure of human hypopharyngeal cells to acidic bile deregulates cancer-related miRNA markers that can be reversed by BAY 11-7082, a pharmacologic NF-κB inhibitor. We repetitively exposed normal human hypopharyngeal primary cells and human hypopharyngeal keratinocytes to bile fluid (400 μmol/L), at pH 4.0 and 7.0, with/without BAY 11-7082 (20 μmol/L). We centred our study on the transcriptional activation of oncogenic miR-21, miR-155, miR-192, miR-34a, miR-375, miR-451a and NF-κB-related genes, previously linked to acidic bile-induced pre-neoplastic events. Our novel findings in vitro are consistent with our hypothesis demonstrating that BAY 11-7082 significantly reverses the acidic bile-induced oncogenic miRNA phenotype, in normal hypopharyngeal cells. BAY 11-7082 strongly inhibits the acidic bile-induced up-regulation of miR-192 and down-regulation of miR-451a and significantly decreases the miR-21/375 ratios, previously related to poor prognosis in hypopharyngeal cancer. This is the first in vitro report that NF-κB inhibition reverses acidic bile-induced miR-21, miR-155, miR-192, miR-34a, miR-375 and miR-451a deregulations in normal human hypopharyngeal cells, suggesting that acidic bile-induced events are directly or indirectly dependent on NF-κB signalling.
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http://dx.doi.org/10.1111/jcmm.13591DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5908126PMC
May 2018

Inhibition of NF-B prevents the acidic bile-induced oncogenic mRNA phenotype, in human hypopharyngeal cells.

Oncotarget 2018 Jan 12;9(5):5876-5891. Epub 2017 Dec 12.

The Yale Larynx Laboratory, Department of Surgery, Yale School of Medicine, New Haven, CT, USA.

Bile-containing gastro-duodenal reflux has been clinically considered an independent risk factor in hypopharyngeal carcinogenesis. We recently showed that the chronic effect of acidic bile, at pH 4.0, selectively induces NF-B activation and accelerates the transcriptional levels of genes, linked to head and neck cancer, in normal hypopharyngeal epithelial cells. Here, we hypothesize that NF-B inhibition is capable of preventing the acidic bile-induced and cancer-related mRNA phenotype, in treated normal human hypopharyngeal cells. In this setting we used BAY 11-7082, a specific and well documented pharmacologic inhibitor of NF-B, and we observed that BAY 11-7082 effectively inhibits the acidic bile-induced gene expression profiling of the NF-B signaling pathway (down-regulation of 72 out of 84 analyzed genes). NF-B inhibition significantly prevents the acidic bile-induced transcriptional activation of NF-B transcriptional factors, RELA (p65) and c-REL, as well as genes related to and commonly found in established HNSCC cell lines. These include anti-apoptotic bcl-2, oncogenic STAT3, EGFR, ∆Np63, TNF-α and WNT5A, as well as cytokines IL-1β and IL-6. Our findings are consistent with our hypothesis demonstrating that NF-B inhibition effectively prevents the acidic bile-induced cancer-related mRNA phenotype in normal human hypopharyngeal epithelial cells supporting an understanding that NF-B may be a critical link between acidic bile and early preneoplastic events in this setting.
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http://dx.doi.org/10.18632/oncotarget.23143DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5814181PMC
January 2018

Mismatch DNA repair mRNA expression profiles in oral melanin pigmentation lesion and hamartomatous polyp of a child with Peutz-Jeghers syndrome.

Pediatr Blood Cancer 2013 Oct 15;60(10):E116-7. Epub 2013 May 15.

Department of Pathology, Medical School, University of Thessaly, Larissa, Greece.

Mismatch DNA repair (MMR) mRNA expression analysis was performed on a biopsy of oral mucosa melanin pigmentation lesion, a hamartomatous polyp and peripheral blood derived from a 12-year-old child with Peutz-Jeghers Syndrome (PJS). We present a deficient MMR system, in a PJS patient, which demonstrated low mRNA levels of hMSH6 and hPMS2 and an increasing MMR deficiency from the non-dysplastic lesion to hamartomatous polyp of PJS with a high risk of cancer.
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http://dx.doi.org/10.1002/pbc.24579DOI Listing
October 2013

Mismatch repair hMSH2, hMLH1, hMSH6 and hPMS2 mRNA expression profiles in precancerous and cancerous urothelium.

Oncol Lett 2013 Jan 19;5(1):283-294. Epub 2012 Oct 19.

Department of Pathology, Medical School, University of Thessaly, Larissa, Thessaly 41110;

Changes in the expression of the mismatch repair (MMR) genes hMSH2, hMLH1, hMSH6 and hPMS2 reflect dysfunction of the DNA repair system that may allow the malignant transformation of tissue cells. The aim of the present study was to address the mRNA expression profiles of the mismatch DNA repair system in cancerous and precancerous urothelium. This is the first study to quantify MMR mRNA expression by applying quantitative real-time PCR (qPCR) and translate the results to mRNA phenotypic profiles (r, reduced; R, regular or elevated) in bladder tumors [24 urothelial cell carcinomas (UCCs) and 1 papillary urothelial neoplasm of low malignant potential (PUNLMP)] paired with their adjacent normal tissues (ANTs). Genetic instability analysis was applied at polymorphic sites distal or close to the hMSH2 and hMLH1 locus. Presenting our data, reduced hMSH2, hMSH6 and hPMS2 mRNA expression profiles were observed in cancerous and precancerous urothelia. Significantly, the ANTs of UCCs revealed the highest percentages of reduced hMSH2 (r(2)), hMSH6 (r(6)) and hPMS2 (p(2)) mRNA phenotypes relative to their tumors (P<0.03). In particular, combined r(2)r(6) (P<0.02) presented a greater difference between ANTs of low-grade UCCs vs. their tumors compared with ANTs of high-grade UCCs (P= 0.000). Reduced hMLH1 (r(1)) phenotype was not expressed in precancerous or cancerous urothelia. The hMSH6 mRNA was the most changed in UCCs (47.8%), while hMSH2, hMLH1 and hPMS2 showed overexpression (47.8, 35 and 30%, respectively) that was associated with gender and histological tumor grading or staging. Genetic instability was rare in polymorphic regions distal to hMLH1. Our data reveal a previously unrecognized hMSH2 and hMSH6 mRNA combined phenotype (r(2)r(6)) correlated with a precancerous urothelium and show that hMLH1 is transcriptionally activated in precancerous or cancerous urothelium. In the present study, it is demonstrated that reduction of hMSH6 mRNA is a frequent event in bladder tumorigenesis and reflects a common mechanism of suppression with hMSH2, while alterations of hMSH2 or hMLH1 mRNA expression in UCCs does not correlate with the allelic imbalance of polymorphic regions harboring the genes.
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http://dx.doi.org/10.3892/ol.2012.979DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3525358PMC
January 2013

hMSH2 and hMLH1 gene expression patterns differ between lung adenocarcinoma and squamous cell carcinoma: correlation with patient survival and response to adjuvant chemotherapy treatment.

Int J Biol Markers 2013 Jan 27;27(4):e400-4. Epub 2013 Jan 27.

Laboratory of Molecular Histopathology, Department of Pathology, University Hospital of Larissa, Larissa, Greece.

Background: We recently showed that the mRNA levels of mismatch repair (MMR) proteins in non-small cell lung carcinoma (NSCLC) tissue specimens and the phenotypic translation of molecular MMR data refines the biology of the MMR system with consequent diagnostic implications in the clinical assessment of lung cancer patients.

Methods: hMLH1 and hMSH2 mRNA expression was previously evaluated by qPCR for 29 NSCLC patients (13 with squamous cell carcinoma [SQC] and 16 with adenocarcinoma [ADC]) and MMR mRNA levels were converted into clinically distinct phenotypic entities. In this study, we evaluated the correlation of the hMSH2 and hMLH1 mRNA phenotypes with patient survival and their response to adjuvant chemotherapy.

Results: hMSH2 and hMLH1 mRNA phenotypic distribution differed between SQC and ADC. The MMR phenotypes differed also between advanced and early stage SQC. SQC patients with an increased hMSH2 expression had a better outcome than patients with a reduced hMSH2 expression. However, ADC patients with an increased hMSH2 expression had a poor outcome compared to those with low hMSH2 levels. SQC patients with a high hMSH2 expression exhibited a better response to adjuvant chemotherapy, whereas ADC patients with high hMSH2 levels had a poor response. ADC patients with low hMSH2 levels showed good response to adjuvant chemotherapy compared to SQC patients bearing the same phenotypic profile.

Conclusions: Our findings show that MMR mRNA phenotypes may be added to the known biological differences between SQC and ADC. hMLH1 and hMSH2 phenotypes distributed differently according to the NSCLC stage. Distinct MMR mRNA phenotypes in SQC and ADC corresponded to patient response to adjuvant chemotherapy.
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http://dx.doi.org/10.5301/JBM.2012.9420DOI Listing
January 2013
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