Screening, identification and experimental design to optimization of the selenite bioremediation by new isolated Bacillus sp. Selena 3 in water.

Background and Objectives: Heavy metals pollution is one of the most important concerns in the world. Selenium is one of the most important elements for the life, but if the absorption of this element in cells increases, it acts as a toxic element. Materials and Methods: In this study, bacterial isolates were screened and isolated from selenium-contaminated soil and water. Twenty-five out of 42 isolates were able to reduce Selenite. Also, the response surface method (RSM) was used to evaluate and optimize the biological reduction of selenite by Selena 3. Factors of bacterial inoculation percentage, time, and amount of selenium oxyanion salt concentration were studied at five levels of -α, -1, 0, +1, and +α. Results: Bacillus sp. Selena 3 was able to reduce 80 mM sodium selenite in less than 4 hours compared to other bacterial isolates. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of sodium selenite Bacillus sp. Selena 3 was reported as 160 and 320 mM, respectively. The results showed that with increasing duration, the percentage of selenite reduction by bacteria increases and the percentage of bacterial inoculation does not have much effect on its reduction. Conclusion: Due to the ability of Bacillus sp. Selena 3 for rapid reduction in significant concentration of selenium oxyanion (SeO32-), this bacterium can be used as an efficient candidate in removing selenite from the environment.

Relative effect of Malayer Shahani and Asgari grapes seed extract on inducing apoptosis in human leukemia cells.

BACKGROUND: Previous studies have suggested that consuming fruit and vegetable can lower the risk of several cancers, including breast, colorectal, and lung cancers. AIMS: The present study aims to investigate the in vitro anticancer effects of Shahani and Asgari grape seed extract (GSE) grown in Malayer City of Iran on HL-60 cancer. However, to the best of the author's knowledge, it is the first time in this study that the antiproliferative effect of Shahani and Asgari GSE is compared. MATERIALS AND METHODS: Shahani and Asgari GSE Was extraction white method of Liquid/liquid extraction with ethyl acetate. Then assessing cytotoxic activities of Shahani and Asgari GSE on the HL-60 cells was tested using MTT assay. RESULTS: The results show that compared with the control group, seed extract of both Shahani and Asgari at the various concentrations (25, 50, 100, and 200 µg/ml) had a significantly inhibitory effect on HL-60 cell proliferation that was dose dependent. However, Shahani GSE at different concentrations (50, 100, and 200 µg/ml) indicated a significantly higher inhibitory effect compared to Asgari GSE. In addition, GSE can induce cell cycle arrest at G0/G1 cells. Furthermore, GSE of Asgari and Shahani remarkably increased the induction of HL-60 cell apoptosis depending on its dose. However, at the concentration of 200 µg/ml, GSE induced cell necrosis rather than apoptosis. CONCLUSION: Seed extract of both Shahani and Asgari at the various concentrations had a significantly inhibitory effect on HL-60 cell proliferation that was dose dependent.

Esophagogastric junction outflow obstruction.

In the Chicago Classification version 4.0 (CCv4), esophagogastric junction outflow obstruction (EGJOO) is manometrically defined as an elevated median integrated relaxation pressure (IRP) and elevated intrabolus pressure (IBP) during supine wet swallows, and persistently elevated median IRP in the upright position. A clinically relevant conclusive diagnosis of EGJOO requires a manometric diagnosis of EGJOO and associated symptoms such as dysphagia and/or chest pain with at least one of the following supportive investigations (pharmacologic provocation, timed barium esophagogram, and/or endoflip). The Chicago Classification is intended for diagnosis of primary esophageal motor disorders, and thus history and endoscopic evaluation are important to exclude conditions (eg, previous surgery, strictures, or masses) that can secondarily generate the EGJOO pattern on HRM. While a manometric finding of EGJOO is often made and can be an early sign of achalasia, more often it is a manometric finding without clinical implications. The proposed changes in CC4.0 have attempted to make the diagnosis more specific, in order to reduce the number of clinically irrelevant diagnoses and avoid confusion by patients and physicians alike.

Esophageal motility disorders on high-resolution manometry: Chicago classification version 4.0©.

Chicago Classification v4.0 (CCv4.0) is the updated classification scheme for esophageal motility disorders using metrics from high-resolution manometry (HRM). Fifty-two diverse international experts separated into seven working subgroups utilized formal validated methodologies over two-years to develop CCv4.0. Key updates in CCv.4.0 consist of a more rigorous and expansive HRM protocol that incorporates supine and upright test positions as well as provocative testing, a refined definition of esophagogastric junction (EGJ) outflow obstruction (EGJOO), more stringent diagnostic criteria for ineffective esophageal motility and description of baseline EGJ metrics. Further, the CCv4.0 sought to define motility disorder diagnoses as conclusive and inconclusive based on associated symptoms, and findings on provocative testing as well as supportive testing with barium esophagram with tablet and/or functional lumen imaging probe. These changes attempt to minimize ambiguity in prior iterations of Chicago Classification and provide more standardized and rigorous criteria for patterns of disorders of peristalsis and obstruction at the EGJ.

Esophageal hypercontractility is abolished by cholinergic blockade.

BACKGROUND: Esophageal hypercontractility (EHC) is considered a major esophageal motor disorder of unclear etiology. Different mechanisms have been proposed, including an imbalance in inhibitory and excitatory esophageal innervation. We hypothesized that patients with EHC suffer from cholinergic hyperactivity. AIM: To interrogate the excitatory and inhibitory neurotransmission in EHC by assessing the esophageal motor response to atropine (ATR) and cholecystokinin (CCK), respectively, in EHC patients. METHOD: We retrospectively reviewed patients who underwent high-resolution manometry (HRM) with pharmacologic challenge in a tertiary referral center between 2007 and 2017. We identified 49 EHC patients who were categorized based on frequency of hypercontractile peristaltic sequence into "frequent" and "infrequent" and motility diagnosis groups. Deglutitive pressure metrics and esophageal motor responses to ATR (12 mcg/kg iv) and CCK (40 ng/kg iv) were analyzed across groups. RESULTS: Atropine abolished hypercontractility across all groups studied, converting nearly half of patients to a motor pattern of ineffective esophageal motility. Abnormal CCK responses primarily occurred in the patient groups with concomitant outflow obstruction. CONCLUSIONS: Hypercontractility is cholinergically mediated in all esophageal motor disorders. Most patients with isolated EHC appear to have excessive cholinergic drive, rather than loss of inhibitory innervation, and might be candidates for treatment with anticholinergic agents.

Protective effect of vitamin E on sperm parameters in rats infected with Candida albicans.

Candida albicans is one of the most frequent pathogens present in the reproductive system. The negative in vitro effects of C. albicans on sperm functions have previously been studied. The current study was undertaken to investigate the effects of C. albicans infection in vivo on sperm quality and to evaluate the efficacy of vitamin E administration in rats infected with C. albicans. In this study, 5 days after infection induction, animals were treated with vitamin E for 5 weeks. Thereafter, sperm parameters, lipid peroxidation (LPO), total antioxidant capacity (TAC), hormonal analysis and testis histology were evaluated. Based on the results, sperm parameters and TAC significantly reduced, while LPO and tissue damage increased (p ≤ .05) following the infection. Hormone analysis showed low LH and testosterone levels in serum of the infected rats. Treatment with vitamin E significantly (p ≤ .05) improved sperm quality and testis histology, increased TAC and reduced LPO. In addition, vitamin E administration significantly increased (p ≤ .05) serum LH and testosterone levels. These results clearly indicate that vitamin E is effective in attenuating the adverse effects of C. albicans infection on male fertility and could be used as a complementary treatment for patients who suffer from fertility disorders following C. albicans infection.

Diagnostic differences in the pharmacologic response to cholecystokinin and amyl nitrite in patients with absent contractility vs type I Achalasia.

BACKGROUND: Absent esophageal contractility (AC) is distinguished from type 1 achalasia (ACH1) during high-resolution manometry (HRM) on the basis of normal or elevated deglutitive integrated relaxation pressure (IRP) values. However, IRP measurements are subject to pressure recording error. We hypothesized that distinctive responses to pharmacologic provocation using amyl nitrite (AN) and cholecystokinin (CCK) could reliably distinguish AC patients from those with ACH1. AIM: To compare esophageal response with AN and CCK in a well-defined cohort of ACH1 and AC patients. METHOD: All available clinical, radiographic, endoscopic, and manometric information in 34 patients with aperistalsis was reviewed to determine the final diagnosis of ACH1 and AC. The differences in response to provocative challenges with the rapid drink challenge (RDC) test and administration of AN and CCK were compared between these two groups. RESULTS: Eighteen patients were diagnosed with ACH1 and sixteen with AC. While IRP values were significantly higher in ACH1, the standard criterion value misclassified four AC patients as having ACH1 and five ACH1 patients as having AC. IRP values on the RDC did not accurately segregate AC from ACH1, but we were able to identify AN and CCK esophageal motor response criteria that allowed correct classification of ACH1 and AC patients. CONCLUSIONS: Nearly a quarter of AC and ACH1 patients may be misdiagnosed based on manometric IRP criteria alone. Differences in the esophageal motor responses to AN and CCK have the potential to facilitate the correct diagnosis in these challenging patients.

Motility Patterns Following Esophageal Pharmacologic Provocation With Amyl Nitrite or Cholecystokinin During High-Resolution Manometry Distinguish Idiopathic vs Opioid-Induced Type 3 Achalasia.

BACKGROUND & AIMS: In some patients, the type 3 achalasia (A3) motor pattern may be an effect of chronic use of high-dose opioids. No motor findings have been identified to differentiate opioid-induced A3 (OA3) from idiopathic A3 (IA3). We investigated whether OA3 could be distinguished from IA3 on the basis of differences in esophageal motor responses to amyl nitrite, cholecystokinin, or atropine. METHODS: We performed a retrospective study of patients who received pharmacologic provocation during esophageal high-resolution manometry from 2007 through 2017 at a tertiary referral center. We identified 26 patients with IA3 (9 women; mean age, 68 ± 13 years) and 24 patients with OA3 (15 women; mean age, 59 ± 10 years). We compared pressure topography metrics during deglutition and after administration of amyl nitrite, cholecystokinin, or atropine between patients with OA3 vs IA3. RESULTS: Amyl nitrite induced a similar relaxation response in both groups, but the rebound contraction of the lower esophageal sphincter during amyl nitrite recovery, and the paradoxical esophageal contraction during the first phase of cholecystokinin response, were both significantly attenuated in patients with OA3. The second phase of cholecystokinin response in patients with OA3 was 100% relaxation, when present, in contrast to only 26% of patients with IA3. There was no significant difference between groups in inhibition of lower esophageal sphincter tone or esophageal body contractility by cholinergic receptor blockade. CONCLUSIONS: Nearly half of patients with an A3 pattern of dysmotility are chronic, daily users of opioids with manometry patterns indistinguishable from those of patients with IA3. Patients with OA3 differ from patients with IA3 in responses to amyl nitrite and cholecystokinin. These findings might be used to identify patients with dysmotility resulting from opioid use.

Pharmacologic interrogation of patients with esophagogastric junction outflow obstruction using amyl nitrite.

BACKGROUND: The Chicago Classification of esophageal motility includes a group of patients who show evidence of esophagogastric junction outflow obstruction (EGJOO) as demonstrated by elevated integrated relaxation pressure (IRP) and preserved peristalsis. Our aim is to classify EGJOO patients based on response to amyl nitrite (AN) during high-resolution manometry. METHODS: Patients were considered to have true EGJOO if elevated IRP during supine swallow persisted in the upright position and was associated with high intrabolus pressure. The EGJ response to AN was compared between patients with achalasia type 2 (A2) and normal esophageal motility. Based on the relaxation gain (deglutitive IRP-AN IRP) value that best discriminated these two groups (10 mm Hg), patients with true EGJOO were categorized as being in either the AN-responsive (AN-R) or AN-unresponsive (AN-U) subgroups. KEY RESULTS: In the group of 49 patients with true EGJOO, the AN response classified 27 patients (IRP = 25 ± 10 mm Hg) with AN-R and 22 patients (IRP = 20 ± 5 mm Hg) with AN-U (P = 0.2). In AN-R, AN produced a relaxation gain and rebound after-contraction response at the EGJ comparable to A2 patients. AN-U patients had an elevated IRP after AN and a relaxation gain similar to normal esophageal motility patients. AN-U patients were obese and had higher prevalence of sleep apnea (P < 0.05). CONCLUSIONS: Among patients with true EGJOO, only half have pharmacologic evidence of impaired LES relaxation. Pharmacologic interrogation of the EGJ is thus necessary to identify the subgroup of EGJOO patients who could be expected to benefit from LES ablative therapies.

Chronic daily opioid exposure is associated with dysphagia, esophageal outflow obstruction, and disordered peristalsis.

BACKGROUND: Opioid receptors are present in the esophagus, and chronic opioid therapy may be associated with esophageal dysfunction. Given the current opioid epidemic in the United States, the potential contribution of opioids to esophageal dysmotility is important from both public health and patient care perspectives. Therefore our aim is to investigate the potential contribution of opioids to dysphagia and the prevalence of major motor disorders in patients undergoing manometric evaluation. METHODS: The anonymized electronic medical records of patients linked to their de-identified high-resolution manometry (HRM) studies were reviewed. The patients were grouped based on their opioid exposure history at the time of HRM: opioid-naïve and chronic daily users. The oral morphine milligram equivalent daily dose (MMED) of opioids was computed. KEY RESULTS: 10% of patients referred for esophageal HRM were taking opioid analgesics on a chronic daily basis, and they had a significantly higher prevalence of dysphagia than their opioid-naïve counterparts. The chronic daily opioid users displayed a significantly higher prevalence of achalasia type 3 (ACH3) and esophagogastric junction outflow obstruction (EGJOO) motility phenotypes. The MMED of opioids was a significant predictor of esophageal pressure metrics and motility diagnoses (P < 0.0001). CONCLUSIONS: Chronic daily opioid intake is associated with impaired deglutitive LES relaxation and disorganized peristaltic sequence. While a minority of patients on chronic daily opioid therapy present with major esophageal motor disorders, they comprise nearly half of ACH3 and a third of EGJOO motility phenotypes.

Cholecystokinin induces esophageal longitudinal muscle contraction and transient lower esophageal sphincter relaxation in healthy humans.

Cholecystokinin (CCK) is known to cause lower esophageal sphincter (LES) relaxation through the activation of inhibitory motor neurons. CCK receptor agonists increase the frequency of transient LES relaxation through a peripheral mechanism. Recent studies show that the longitudinal muscle contraction (LMC)-related axial stretch might play a role in the LES relaxation by activating the mechanosensitive inhibitory motor neurons. The aim of our study was to determine whether the CCK-induced LES relaxation and the characteristics of LMC resemble those seen with spontaneous transient LES relaxation in humans. Nine healthy volunteers (5 Fr, 40 ± 12 yr) received escalating doses of CCK-octapeptide (CCK-8) (5, 10, 20, and 40 ng/kg). All subjects demonstrated a monophasic response to 5 ng/kg of CCK-8. In the majority of subjects, this response consisted of partial LES relaxation. All subjects showed a biphasic response to 40 ng/kg of CCK-8. The latter in most subjects consisted of 1) a period of partial relaxation followed by 2) complete LES relaxation along with crural diaphragm inhibition. The length of the esophagus decreased by 0.9 ± 0.4 cm, and muscle thickness increased by 40 ± 14% to 1.4 ± 0.2 mm ( P < 0.05) during initial partial LES relaxation. During complete LES relaxation there was greater LMC, as demonstrated by an esophageal shortening of 1.9 ± 0.5 cm and an increase in muscle thickness of 100 ± 16% ( P < 0.01). The complete phase 2 LES relaxation typically terminated with a robust after-contraction. Atropine significantly attenuated the CCK-induced esophageal LMC, prevented crural diaphragm inhibition, and abolished the phase 2 complete LES relaxation. NEW & NOTEWORTHY The phenotypic features of CCK-induced longitudinal muscle contraction (LMC), complete lower esophageal sphincter (LES) relaxation, and crural diaphragm inhibition, followed by a robust after-contraction, resemble those seen during spontaneous transient LES relaxation. A strong temporal relationship between the LMC and complete transient LES relaxation supports our hypothesis that the LMC plays an important role in the LES relaxation and crural diaphragmatic inhibition.

Impaired upper esophageal sphincter reflexes in patients with supraesophageal reflux disease.

BACKGROUND & AIMS: Normal responses of the upper esophageal sphincter (UES) and esophageal body to liquid reflux events prevent esophagopharyngeal reflux and its complications, however, abnormal responses have not been characterized. We investigated whether patients with supraesophageal reflux disease (SERD) have impaired UES and esophageal body responses to simulated reflux events. METHODS: We performed a prospective study of 25 patients with SERD (age, 19-82 y; 13 women) and complaints of regurgitation and supraesophageal manifestations of reflux. We also included 10 patients with gastroesophageal reflux disease (GERD; age, 32-60 y; 7 women) without troublesome regurgitation and supraesophageal symptoms and 24 healthy asymptomatic individuals (controls: age, 19-49 y; 13 women). UES and esophageal body pressure responses, along with luminal distribution of infusate during esophageal rapid and slow infusion of air or liquid, were monitored by concurrent high-resolution manometry and intraluminal impedance. RESULTS: A significantly smaller proportion of patients with SERD had UES contractile reflexes in response to slow esophageal infusion of acid than controls or patients with GERD. Only patients with SERD had abnormal UES relaxation responses to rapid distension with saline. Diminished esophageal peristaltic contractions resulted in esophageal stasis in patients with GERD or SERD. CONCLUSIONS: Patients with SERD and complaints of regurgitation have impaired UES and esophageal responses to simulated liquid reflux events. These patterns could predispose them to esophagopharyngeal reflux.

Prevention of esophagopharyngeal reflux by augmenting the upper esophageal sphincter pressure barrier.

OBJECTIVES/HYPOTHESIS: Incompetence of the upper esophageal sphincter (UES) is fundamental to the occurrence of esophagopharyngeal reflux (EPR), and development of supraesophageal manifestations of reflux disease (SERD). However, therapeutic approaches to SERD have not been directed to strengthening of the UES barrier function. Our aims were to demonstrate that EPR events can be experimentally induced in SERD patients and not in healthy controls, and ascertain if these events can be prevented by application of a modest external cricoid pressure. STUDY DESIGN: Individual case control study. METHODS: We studied 14 SERD patients (57 ± 13 years, 8 females) and 12 healthy controls (26 ± 3 years, 7 females) by concurrent intraesophageal slow infusion and pharyngoscopic and manometric technique without and with the application of a sustained predetermined cricoid pressure to induce, detect, and prevent EPR, respectively. RESULTS: Slow esophageal infusion (1 mL/s) of 60 mL of HCl resulted in a total of 16 objectively confirmed EPR events in none patients and none in healthy controls. All patients developed subjective sensation of regurgitation. Sustained cricoid pressure resulted in a significant UES pressure augmentation in all participants. During application of sustained cricoid pressure, slow intraesophageal infusion resulted in only one EPR event (P < .01). CONCLUSIONS: Slow esophageal liquid infusion unmasks UES incompetence evidenced as the occurrence of EPR. Application of 20 to 30 mm Hg cricoid pressure significantly increases the UES intraluminal pressure and prevents pharyngeal reflux induced by esophageal slow liquid infusion. These techniques can be useful in diagnosis and management of UES incompetence in patients suffering from supraesophageal manifestations of reflux disease.

Role of peripheral reflexes in the initiation of the esophageal phase of swallowing.

The aim of this study was to determine the role of peripheral reflexes in initiation of the esophageal phase of swallowing. In 10 decerebrate cats, we recorded electromyographic responses from the pharynx, larynx, and esophagus and manometric data from the esophagus. Water (1-5 ml) was injected into the nasopharynx to stimulate swallowing, and the timing of the pharyngeal and esophageal phases of swallowing was quantified. The effects of transection or stimulation of nerves innervating the esophagus on swallowing and esophageal motility were tested. We found that the percent occurrence of the esophageal phase was significantly related to the bolus size. While the time delays between the pharyngeal and esophageal phases of swallowing were not related to the bolus size, they were significantly more variable than the time delays between activation of muscles within the pharyngeal phase. Transection of the sensory innervation of the proximal cervical esophagus blocked or significantly inhibited activation of the esophageal phase in the proximal cervical esophagus. Peripheral electrical stimulation of the pharyngoesophageal nerve activated the proximal cervical esophagus, peripheral electrical stimulation of the vagus nerve activated the distal cervical esophagus, and peripheral electrical stimulation the superior laryngeal nerve (SLN) had no effect on the esophagus. Centripetal electrical stimulation of the SLN activated the cervical component of the esophageal phase of swallowing before initiation of the pharyngeal phase. Therefore, we concluded that initiation of the esophageal phase of swallowing depends on feedback from peripheral reflexes acting through the SLN, rather than a central program.

Functional connectivity of the cortical swallowing network in humans.

INTRODUCTION: Coherent fluctuations of blood oxygenation level dependent (BOLD) signal have been referred to as "functional connectivity" (FC). Our aim was to systematically characterize FC of underlying neural network involved in swallowing, and to evaluate its reproducibility and modulation during rest or task performance. METHODS: Activated seed regions within known areas of the cortical swallowing network (CSN) were independently identified in 16 healthy volunteers. Subjects swallowed using a paradigm driven protocol, and the data analyzed using an event-related technique. Then, in the same 16 volunteers, resting and active state data were obtained for 540 s in three conditions: 1) swallowing task; 2) control visual task; and 3) resting state; all scans were performed twice. Data was preprocessed according to standard FC pipeline. We determined the correlation coefficient values of member regions of the CSN across the three aforementioned conditions and compared between two sessions using linear regression. Average FC matrices across conditions were then compared. RESULTS: Swallow activated twenty-two positive BOLD and eighteen negative BOLD regions distributed bilaterally within cingulate, insula, sensorimotor cortex, prefrontal and parietal cortices. We found that: 1) Positive BOLD regions were highly connected to each other during all test conditions while negative BOLD regions were tightly connected among themselves; 2) Positive and negative BOLD regions were anti-correlated at rest and during task performance; 3) Across all three test conditions, FC among the regions was reproducible (r>0.96, p<10(-5)); and 4) The FC of sensorimotor region to other regions of the CSN increased during swallowing scan. CONCLUSIONS: 1) Swallow activated cortical substrates maintain a consistent pattern of functional connectivity; 2) FC of sensorimotor region is significantly higher during swallow scan than that observed during a non-swallow visual task or at rest.

Reproducibility of swallow-induced cortical BOLD positive and negative fMRI activity.

Functional MRI (fMRI) studies have demonstrated that a number of brain regions (cingulate, insula, prefrontal, and sensory/motor cortices) display blood oxygen level-dependent (BOLD) positive activity during swallow. Negative BOLD activations and reproducibility of these activations have not been systematically studied. The aim of our study was to investigate the reproducibility of swallow-related cortical positive and negative BOLD activity across different fMRI sessions. We studied 16 healthy volunteers utilizing an fMRI event-related analysis. Individual analysis using a general linear model was used to remove undesirable signal changes correlated with motion, white matter, and cerebrospinal fluid. The group analysis used a mixed-effects multilevel model to identify active cortical regions. The volume and magnitude of a BOLD signal within each cluster was compared between the two study sessions. All subjects showed significant clustered BOLD activity within the known areas of cortical swallowing network across both sessions. The cross-correlation coefficient of percent fMRI signal change and the number of activated voxels across both positive and negative BOLD networks were similar between the two studies (r ≥ 0.87, P < 0.0001). Swallow-associated negative BOLD activity was comparable to the well-defined "default-mode" network, and positive BOLD activity had noticeable overlap with the previously described "task-positive" network. Swallow activates two parallel cortical networks. These include a positive and a negative BOLD network, respectively, correlated and anticorrelated with swallow stimulus. Group cortical activity maps, as well as extent and amplitude of activity induced by volitional swallowing in the cortical swallowing network, are reproducible between study sessions.

Response of the upper esophageal sphincter to esophageal distension is affected by posture, velocity, volume, and composition of the infusate.

BACKGROUND & AIMS: Studies of the pressure response of the upper esophageal sphincter (UES) to simulated or spontaneous gastroesophageal reflux have shown conflicting results. These discrepancies could result from uncontrolled influence of variables such as posture, volume, and velocity of distension. We characterized in humans the effects of these variables on UES pressure response to esophageal distension. METHODS: We studied 12 healthy volunteers (average, 27 ± 5 years old; 6 male) using concurrent esophageal infusion and high-resolution manometry to determine UES, lower esophageal sphincter, and intraesophageal pressure values. Reflux events were simulated by distal esophageal injections of room temperature air and water (5, 10, 20, and 50 mL) in individuals in 3 positions (upright, supine, and semisupine). Frequencies of various UES responses were compared using χ(2) analysis. Multinomial logistical regression analysis was used to identify factors that determine the UES response. RESULTS: UES contraction and relaxation were the overriding responses to esophageal water and air distension, respectively, in a volume-dependent fashion (P < .001). Water-induced UES contraction and air-induced UES relaxation were the predominant responses among individuals in supine and upright positions, respectively (P < .001). The prevalence of their respective predominant response significantly decreased in the opposite position. Proximal esophageal dp/dt significantly and independently differentiated the UES response to infusion with water or air. CONCLUSIONS: The UES response to esophageal distension is affected by combined effects of posture (spatial orientation of the esophagus), physical properties, and volume of refluxate, as well as the magnitude and rate of increase in intraesophageal pressure. The UES response to esophageal distension can be predicted using a model that incorporates these factors.

Production of a recombinant anti-human CD4 single-chain variable-fragment antibody using phage display technology and its expression in Escherichia coli.

Single-chain variable fragment (scFv) is a fusion protein of the variable regions of the heavy (VH) and light (VL) chains of immunoglobulin, connected with a short linker peptide of 10 to about 20 amino acids. In this study, the scFv of a monoclonal antibody against the third domain of human CD4 was cloned from OKT4 hybridoma cells using the phage display technique and produced in E. coli. The expression, production, and purification of anti-CD4 scFv were tested using SDS-PAGE and Western blot, and the specificity of anti-CD4 scFv was examined using ELISA. A 31 kDa recombinant anti-CD4 scFv was expressed and produced in bacteria, which was confirmed by SDS-PAGE and Western blot assays. Sequence analysis proved the ScFv structure of the construct. It was able to bind to CD4 in quality ELISA assay. The canonical structure of anti-CD4 scFv antibody was obtained using the SWISS_MODEL bioinformatics tool for comparing with the scFv general structure. To the best of our knowledge, this is the first report for generating scFv against human CD4 antigen. Engineered anti-CD4 scFv could be used in immunological studies, including fluorochrome conjugation, bispecific antibody production, bifunctional protein synthesis, and other genetic engineering manipulations. Since the binding site of our product is domain 3 (D3) of the CD4 molecule and different from the CD4 immunological main domain, including D1 and D2, further studies are needed to evaluate the anti-CD4 scFv potential for diagnostic and therapeutic applications.

Restricted leptin antagonism as a therapeutic approach to treatment of autoimmune diseases.

Leptin, the adipocyte derived hormone, has a pivotal role in regulating energy homeostasis and appetite. Beyond this essential role in bodyweight control, leptin also regulates the immune responses. Leptin has pro-inflammatory effects on T cell populations, shifting the T helper balance towards a TH1 phenotype, through induction of pro-inflammatory cytokines and stimulation of macrophage and natural killer cell function. Acute starvation reduces serum leptin levels, resulting in an impaired cellular immune response. The TH1 pro-inflammatory immune response, a homeostatic response mediated by the low leptin levels, is also impaired during starvation. Leptin-deficient or leptin receptor mutant mice are protected against the development of several inflammatory or various T cell-dependent autoimmune diseases. Therefore, leptin appears to have a central role in the immune response and low leptin levels may protect against autoimmune disease. Here we review the role of leptin in the immune responses, with emphasis on autoimmune diseases. We will also discuss the application of leptin antagonist therapy for prevention and treatment of immunity related disorders.