The Natural History of Eosinophilic Gastrointestinal Diseases Is Influenced by Age of Onset and Location of Involvement.

INTRODUCTION: It is unknown whether concomitant esophageal involvement or anatomic location of eosinophilic infiltration affects the natural history of eosinophilic gastrointestinal disease (EGID). METHODS: A retrospective cohort study was performed using the University of North Carolina EGID Clinicopathologic Database. Patients were adults and children with a prior EGID diagnosis based on clinicopathologic features. Demographics, clinical characteristics, treatment information, and procedural data were extracted from medical records. Clinical course and flare history were characterized. RESULTS: Among 97 patients, 43% had EGID + esophageal involvement and 57% had EGID only. Patients with esophageal involvement had a longer diagnostic delay preceding diagnosis (36.6 vs 11.6 months, P = 0.001), more dysphagia (50% vs 18%; P = 0.001), required more chronic therapy (77% vs 52%, P = 0.016), and exhibited more progressive disease (25% vs 6%, P = 0.027). A continuous disease course was most common in eosinophilic gastritis (78%) while patients with eosinophilic gastritis + eosinophilic enteritis (29%) and eosinophilic enteritis + eosinophilic colitis (50%) had the highest proportion of progressive and relapsing disease, respectively ( P = 0.045). A continuous disease course occurred more frequently in children (71%, P = 0.03) and those with single organ involvement (65%), whereas adults had more relapsing (39%) or progressive disease (18%). DISCUSSION: EGIDs with and without esophageal involvement display many similarities, although patients with esophageal involvement more frequently had dysphagia, had progressive disease courses, and required more chronic therapy. Location of involvement and age of onset affected the natural history with higher proportions of relapsing or progressive disease seen in adults and patients with small bowel or multiorgan involvement while a continuous disease course was more common in children and patients with gastric-only involvement.

Clinical Features and Treatment Response to Topical Steroids in Ethnic and Racial Minority Patients With Eosinophilic Esophagitis.

INTRODUCTION: Differences in eosinophilic esophagitis (EoE) presentation and outcomes by ethnicity or race remain understudied. We aimed to determine whether EoE patients of Hispanic/Latinx ethnicity or non-White race have differences in presentation at diagnosis or response to topical corticosteroid (tCS) treatment. METHODS: This retrospective cohort study included subjects of any age with a new diagnosis of EoE and documentation of ethnicity or race. For those who had treatment with tCS and follow-up endoscopy/biopsy, we assessed histologic response (<15 eosinophils/hpf), global symptom response, and endoscopic response. Hispanic EoE patients were compared with non-Hispanics at baseline and before and after treatment. The same analyses were repeated for White vs non-Whites. RESULTS: Of 1,026 EoE patients with ethnicity data, just 23 (2%) were Hispanic. Most clinical features at presentation were similar to non-Hispanic EoE patients but histologic response to tCS was numerically lower (38% vs 57%). Non-White EoE patients (13%) were younger at diagnosis and had less insurance, lower zip code-level income, shorter symptom duration, more vomiting, less dysphagia and food impaction, fewer typical endoscopic features, and less dilation. Of 475 patients with race data treated with tCS, non-Whites had a significantly lower histologic response rate (41% vs 59%; P = 0.01), and odds of histologic response remained lower after controlling for potential confounders (adjusted odds ratio 0.40, 95% confidence intervals: 0.19-0.87). DISCUSSION: Few EoE patients at our center were Hispanic, and they had similar clinical presentations as non-Hispanics. The non-White EoE group was larger, and presentation was less dysphagia-specific. Non-White patients were also less than half as likely to respond to tCS.

Eosinophilic esophagitis patients with multiple atopic conditions: Clinical characteristics and treatment response to topical steroids.

BACKGROUND: Patients with eosinophilic esophagitis (EoE) typically have concomitant atopic conditions, but whether there are differences in presentation or treatment response by the number of atopic diseases is unknown. OBJECTIVE: To determine whether patients with EoE having multiple atopic conditions have differences in presentation or response to topical corticosteroid (TCS) treatment. METHODS: We performed a retrospective cohort study of adults and children with newly diagnosed EoE. The total number of atopic comorbidities (allergic rhinitis, asthma, eczema, food allergy) was calculated. Patients with at least 2 atopic conditions other than allergic rhinitis were defined as having multiple atopic conditions and their baseline characteristics were compared with those with less than 2 atopic conditions. Histologic, symptom, and endoscopic responses to TCS treatment were also compared with bivariable and multivariable analyses. RESULTS: Of the 1020 patients with EoE having atopic disease information, 235 (23%) had 1 atopic comorbidity, 211 (21%) had 2, 113 (11%) had 3, and 34 (3%) had 4. At baseline, the 180 (18%) patients with 2 or more atopic diseases were younger and had more vomiting, less abdominal pain, more exudates and edema on endoscopy, and higher peak eosinophil counts. Among those treated with TCS, there was a trend toward better global symptom response in patients with less than 2 atopic conditions, but there was no difference in histologic or endoscopic response compared with those with 2 or more atopic conditions. CONCLUSION: There were differences in the initial presentation of EoE between those with and without multiple atopic conditions, but there were no major differences in histologic treatment response to corticosteroids by atopic status.

Higher Body Mass Index Is Associated With Decreased Treatment Response to Topical Steroids in Eosinophilic Esophagitis.

BACKGROUND & AIMS: Understanding which eosinophilic esophagitis (EoE) patients will respond to treatment with topical corticosteroids (tCS) remains challenging, and it is unknown whether obesity impacts treatment response. This study aimed to determine whether treatment outcomes to tCS in EoE patients vary by body mass index (BMI). METHODS: This retrospective cohort study of the University of North Carolina EoE Clinicopathologic database assessed subjects age 14 years or older with a new diagnosis of EoE. Their BMI was calculated and histologic, symptom, and endoscopic responses were recorded after tCS treatment. The treatment response of obese (BMI, ≥30 kg/m2) and nonobese EoE status was compared using bivariate and multivariate analyses. RESULTS: We identified 296 EoE patients treated with tCS. Baseline characteristics were similar, although obese EoE patients had more heartburn and hiatal hernias. Histologic response was higher for those who were nonobese compared with obese at fewer than 15 (61% vs 47%; P = .049) and 6 or fewer (54% vs 38%; P = .02) eosinophils per high-power field, respectively. In addition, nonobese patients had significantly greater endoscopic and symptomatic responses. On multivariate analysis, increasing BMI was associated independently with decreased histologic response after accounting for age, heartburn, dilation, and hiatal hernia whether BMI was assessed as a continuous variable (adjusted odds ratio [aOR], 0.93; 95% CI, 0.89-0.98), as nonobese vs obese (aOR, 0.38; 95% CI, 0.21-0.68), or in 4 categories (overweight vs normal [aOR, 0.46; 95% CI, 0.26-0.84] or obese vs normal [aOR, 0.26; 95% CI, 0.13-0.51]). CONCLUSIONS: As BMI increases in EoE patients, the odds of histologic, symptomatic, and endoscopic responses to tCS decreases, with obese patients having an approximately 40% decrease in odds of response.

Association between eosinophilic esophagitis and esophageal dysmotility: A systematic review and meta-analysis.

BACKGROUND: There is conflicting evidence about the association between eosinophilic esophagitis (EoE) and esophageal motility disorders. The aim of this study was to evaluate esophageal manometry findings in EoE. METHODS: We conducted a systematic review using PubMed, EMBASE, and Web of Science. All articles from 1990 to 2021 with EoE patients who underwent esophageal manometry were eligible. We also included pertinent abstracts from national conferences from 2015 to 2020. The primary outcomes were the prevalence of specific Chicago 3 Classification (CCv3) diagnoses in EoE, as well as broader categories of non-relaxing lower esophageal sphincter, and major and minor peristaltic disorders. When multiple studies reported a specific outcome, we performed random effects meta-analysis to obtain pooled prevalence of each outcome. To reduce heterogeneity, we restricted meta-analysis to high-resolution manometry (HRM) studies only. KEY RESULTS: Of 763 publications identified, 27 original studies met criteria for inclusion, encompassing 706 EoE patients; 14 studies (425 patients) had HRM and underwent meta-analysis. The pooled prevalence of any motility abnormality was 53% (95% CI: 43%-63%), largely comprised of minor motility disorders such as ineffective esophageal motility and fragmented peristalsis. Major motility disorders, classified by CCv3, were less common in EoE, with pooled prevalence of 2% (0%-7%), 10% (5%-16%), and 1% (0%-3%), for achalasia, esophagogastric-junction outflow obstruction, and hypercontractile disorders, respectively. CONCLUSION AND INFERENCES: Non-specific motility disorders were common in patients with EoE, but major motility disorders were rare. Further studies are needed to determine the relationship between eosinophilic infiltration and the clinical relevance of abnormal esophageal motility findings in this population.

Association of eosinophilic esophagitis with autoimmune and connective tissue disorders, and the impact on treatment response.

Eosinophilic esophagitis (EoE) has been associated with autoimmune (AI) and connective tissue disorders (CTDs), but clinical correlates and treatment response to topical corticosteroids (tCS) for patients with both conditions are not well known. We aimed to determine the prevalence and clinical features of AI/CTDs in EoE patients, and assess the response to tCS. In this retrospective cohort study of adults and children newly diagnosed with EoE in the University of North Carolina EoE Clinicopathologic database, we extracted clinical characteristics and treatment response data. We compared EoE patients with and without AI/CTDs, identified independently associated factors, and explored treatment responses. Of 1029 EoE patients, 61 (5.9%) had an AI/CTDs. The most common AI/CTDs were psoriasis/psoriatic arthritis (P/PA) (1.7%), Hashimoto's (1.2%), and rheumatoid arthritis (RA) (1%). Compared to those without AI/CTDs, AI/CTDs patients were older (35 vs. 28 years, P = 0.004), more likely to be female (51% vs. 30%, P = 0.001), have insurance (93% vs. 78%, P = 0.004) and a longer symptom duration prior to EoE diagnosis (10 vs. 7 years, P = 0.02). Older age, female sex, having insurance, and having allergic rhinitis were independently associated with AI/CTDs. AI/CTD patients with EoE were less likely to have a symptom response (47% vs. 79%, P = 0.003). Overlap between EoE and AI/CTDs was uncommon, seen in approximately 6%, with P/PA, Hashimoto's, and RA being most frequent. In conclusion, older age, female sex, having insurance, and allergic rhinitis were independently associated with AI/CTDs. EoE patients with AI/CTDs had less symptom response, with trendtowards lower endoscopic and histologic responses, to tCS therapy.

Loss to Follow-Up and Health Care Utilization After Initial Diagnosis of Eosinophilic Esophagitis.

BACKGROUND: Eosinophilic esophagitis (EoE) is a chronic disease, but the extent of patient loss to follow-up (LTFU) and health care utilization has not been fully investigated. AIM: To determine frequency and predictors of LTFU and health care utilization in EoE patients. METHODS: In this retrospective cohort study, we extracted data from patients with a new diagnosis of EoE. Follow-up time for each patient was calculated as the time from the first diagnostic endoscopy to the last GI-related contact date in the medical record. Patients with and without LTFU were compared, and the volume of EoE-related health care interactions was recorded. RESULTS: Of 944 EoE cases, 249 (26%) met the definition for LTFU. Major reasons for LTFU were never being scheduled (45%) and inability to contact patients (40%). Factors independently associated with regular follow-up were having insurance (aOR 2.89; 95% CI 1.85-4.50), white race (aOR 2.16; 95% CI 1.37-3.41), and longer symptom length (aOR 1.04 per year; 95% CI 1.01-1.08). At the time of last contact, patients with follow-up had better symptom response (55% vs. 12%; p < 0.001), improved esophageal caliber (14.3 vs. 12.4 mm; p = 0.005), and more histologic response (45% vs. 4% at 15 eos/hpf; p < 0.001). Health care utilization was high, with an average of 4.6 endoscopies and 4.0 clinic visits over the follow-up period. CONCLUSIONS: LTFU of newly diagnosed EoE cases was common and associated with lack of insurance, non-white race, and shorter symptom duration. Those who followed up had high health care utilization but improved response rates. Strategies are needed to help decrease LTFU in EoE.

A Gap in Care Leads to Progression of Fibrosis in Eosinophilic Esophagitis Patients.

BACKGROUND & AIMS: There are few data assessing disease progression in eosinophilic esophagitis (EoE) after diagnosis. We aimed to determine outcomes and assess for progression of fibrosis in patients with EoE with a gap in their regular care. METHODS: In this retrospective cohort study of newly diagnosed patients with EoE, a "gap" in care was defined as ≥2 years without medical contact for EoE. For inclusion, a gap in care and both pre- and post-gap endoscopies were required. Patients with and without a gap were compared. Data were also compared in gap patients before the gap and after EoE care resumed, and progression of fibrosis and predictors were assessed. RESULTS: Of 701 patients with EoE, 95 (14%) had a gap in care (mean time without care, 4.8 ± 2.3 years). Post-gap, 12% presented with food impaction requiring emergency evaluation. Compared with pre-gap, patients post-gap had higher endoscopic severity (2.4 vs 1.5; P < .001) and smaller esophageal diameters (11.0 vs 12.7 mm; P = .04). Strictures were more prevalent with longer gap time (P < .05 for trend). Each additional year of gap time increased odds of stricture by 26%, even after accounting for pre-gap dilation. Additionally, of 67 patients without pre-gap fibrosis, 25 (37%) had at least one fibrotic feature (stricture, narrowing, or requiring dilation) post-gap. CONCLUSIONS: A gap in care of ≥2 years in patients with EoE was associated with signs of increased disease activity, and progression to fibrostenosis was noted, particularly with longer gaps in care. Because EoE can progress to fibrosis even after diagnosis, regular care in patients with EoE is required, perhaps at intervals <2 years.

Psychiatric Comorbidities Are Highly Prevalent in Nonesophageal Eosinophilic Gastrointestinal Diseases.

BACKGROUND & AIMS: The prevalence of psychiatric comorbidity in nonesophageal eosinophilic gastrointestinal disorders (EGIDs) has not been studied. We aimed to ascertain the prevalence of psychiatric diagnoses and psychiatric medication use in children, adolescents, and adults with EGIDs and to assess whether psychiatric comorbidity affects clinical presentation. METHODS: This was a retrospective cohort study of newly diagnosed patients with a nonesophageal EGID at the University of North Carolina from 2008 to 2020. Psychiatric diagnoses and medications were extracted from medical records. We compared the clinical and demographic features of EGID patients with and without psychiatric diagnoses. RESULTS: Of 79 patients (mean 23.3 years of age, 53% male, 78% White) with a nonesophageal EGID diagnosis, 40 (51%) were diagnosed with a comorbid psychiatric disease. Anxiety (37%) and depression (28%) were most common. There were also 40 (51%) patients treated medically for a psychiatric diagnosis. Patients with a psychiatric diagnosis were more commonly ≥18 years of age at the time of EGID diagnosis (odds ratio [OR], 3.95, 95% confidence interval [CI], 1.20-13.02) and had endorsed symptoms of nausea (OR, 5.31; 95% CI, 1.33-21.22) and dysphagia (OR, 4.24; 95% CI, 1.18-15.26). CONCLUSION: Psychiatric diagnoses were very common in nonesophageal EGID patients with approximately 7 in 10 adults and one-third of children diagnosed. Similar proportions were found for psychiatric medication use. We also found that psychiatric illness may influence age of clinical presentation and symptoms. Providers should assess for concomitant psychiatric comorbidities in EGID patients.

Older patients with eosinophilic esophagitis have high treatment response to topical steroids.

BACKGROUND: There are few data assessing treatment response in older eosinophilic esophagitis (EoE) patients and we evaluated treatment outcomes to topical corticosteroids (tCS) in this older population. METHODS: This retrospective cohort study of the UNC EoE Clinicopathologic database included subjects with a new diagnosis of EoE treated with tCS. Histologic responses, global symptom response, and endoscopic changes were recorded. Older EoE patients (≥65 years) were compared to younger EoE patients (<65). RESULTS: We identified 467 EoE patients treated with tCS, 12 (3%) of whom were ≥65 years. Compared to those <65 years, patients ≥65 had longer symptom duration and worse endoscopy scores, but most clinical features were similar. Post-treatment peak eosinophil counts trended higher in the <65 group (25.0 vs 5.5; p = 0.07). Histological response was greater in the ≥65 population at <15 eos/hpf (92% vs 57%; p = 0.02), ≤6 eos/hpf (83% vs 50%; p = 0.02), and <1 eos/hpf (58% vs 29%; p = 0.03). Older age was independently associated with increased odds of histologic response (adjusted OR 8.48, 95% CI: 1.08-66.4). CONCLUSIONS: EoE patients ≥65 years had a higher likelihood of responding to tCS therapy, suggesting they should be studied more closely and included in future trials.

Treatment with compounded fluticasone suspension improves the clinical, endoscopic, and histologic features of eosinophilic esophagitis.

No approved medication exists for the treatment of eosinophilic esophagitis (EoE) in the United States, which forces patients to utilize off-label drugs and/or create their own formulations. We assessed the efficacy of a standardized compounded fluticasone suspension. To do this, we performed a retrospective cohort study identifying all EoE patients treated with compounded fluticasone. Compounded fluticasone was prescribed during routine clinical care and dispensed by a specialty compounding pharmacy. Clinical data were extracted from medical records. Outcomes (symptomatic, endoscopic, and histologic) were assessed after the initial and last compounded fluticasone treatment in our system. There were 27 included patients (mean age 34.2; 67% male; 96% white) treated for a mean length of 5.4 ± 4.4 months. The majority (89%) previously utilized dietary elimination or topical corticosteroids, and many (75%) had primary non-response or secondary loss of response to these treatments. After starting compounded fluticasone, symptoms and endoscopic findings improved [dysphagia (89 vs. 56%, P = 0.005), food impaction (59 vs. 4%, P = 0.003), heartburn (26 vs. 4%, P = 0.01), chest pain (26 vs. 8%, P = 0.05), white plaques (63 vs. 32%; P = 0.005), furrows (81 vs. 60%; P = 0.06), and edema (15 vs. 4%; P = 0.16)]. The median of the peak eosinophil counts decreased from 52 to 37 eos/hpf (P = 0.10) and 35% of patients achieved <15 eos/hpf. In conclusion, compounded fluticasone provided a significant improvement in symptoms and endoscopic findings, with more than a third achieving histologic response in a treatment refractory EoE population. Compounded fluticasone should be considered as an EoE management option.

Sodium-hydrogen exchanger regulatory factor-1 (NHERF1) confers salt sensitivity in both male and female models of hypertension in aging.

Hypertension is a risk factor for premature death and roughly 50% of hypertensive patients are salt-sensitive. The incidence of salt-sensitive hypertension increases with age. However, the mechanisms of salt-sensitive hypertension are not well understood. We had demonstrated decreased renal sodium­hydrogen exchanger regulatory factor 1 (NHERF1) expression in old salt-resistant F344 rats. Based on those studies we hypothesized that NHERF1 expression is required for the development of some forms of salt-sensitive hypertension. To address this hypothesis, we measured blood pressure in NHERF1 expressing salt-sensitive 4-mo and 24-mo-old male and female Fischer Brown Norway (FBN) rats male and female 18-mo-old NHERF1 knock-out (NHERF1-/-) mice and wild-type (WT) littermates on C57BL/6J background after feeding high salt (8% NaCl) diet for 7 days. Our data demonstrate that 8% salt diet increased blood pressure in both male and female 24-mo-old FBN rats but not in 4-mo-old FBN rats and in 18-mo-old male and female WT mice but not in NHERF1-/- mice. Renal dopamine 1 receptor (D1R) expression was decreased in 24-mo-old rats, compared with 4-mo-old FBN rats. However, sodium chloride cotransporter (NCC) expression increased in 24-mo-old FBN rats. In FBN rats, age had no effect on NaK ATPase α1 and NKCC2 expression. By contrast, high salt diet increased the renal expressions of NKCC2, and NCC in 24-mo-old FBN rats. High salt diet also increased NKCC2 and NCC expression in WT mice but not NHERF1-/- mice. Our data suggest that renal NHERF1 expression confers salt sensitivity with aging, associated with increased expression of sodium transporters.

The Antiarrhythmic Drug, Amiodarone, Decreases AKT Activity and Sensitizes Human Acute Myeloid Leukemia Cells to Apoptosis by ABT-263.

BACKGROUND: Successful treatment of leukemia requires new medications to combat drug resistance, but the development of novel therapies is an arduous and risky endeavor. Repurposing currently approved drugs or those already in clinical development to treat other indications is a more practical approach. Moreover, combinatorial therapeutics are often more efficacious than single agent therapeutics because the former can simultaneously target multiple pathways that mitigate tumor aggressiveness and induce cancer cell death. MATERIAL AND METHODS: In this study, we combined the class III antiarrhythmic agent amiodarone and the BH3 mimetic ABT-263 based on data from a prior drug screen to assess the degree of apoptotic induction in 2 human leukemia cell lines. RESULTS: The combination yielded statistically significant increases in apoptosis in both cell lines by downregulating AKT activity and increasing cleaved caspase-3. CONCLUSIONS: Overall, our findings suggest that combining K+ channel blockers with prosurvival Bcl-2 family inhibitors is a promising therapeutic approach in treating leukemia.

Loss of NHERF-1 expression prevents dopamine-mediated Na-K-ATPase regulation in renal proximal tubule cells from rat models of hypertension: aged F344 rats and spontaneously hypertensive rats.

Dopamine decreases Na-K-ATPase (NKA) activity by PKC-dependent phosphorylation and endocytosis of the NKA α1. Dopamine-mediated regulation of NKA is impaired in aging and some forms of hypertension. Using opossum (OK) proximal tubule cells (PTCs), we demonstrated that sodium-hydrogen exchanger regulatory factor-1 (NHERF-1) associates with NKA α1 and dopamine-1 receptor (D1R). This association is required for the dopamine-mediated regulation of NKA. In OK cells, dopamine decreases NHERF-1 association with NKA α1 but increases its association with D1R. However, it is not known whether NHERF-1 plays a role in dopamine-mediated NKA regulation in animal models of hypertension. We hypothesized that defective dopamine-mediated regulation of NKA results from the decrease in NHERF-1 expression in rat renal PTCs isolated from animal models of hypertension [spontaneously hypertensive rats (SHRs) and aged F344 rats]. To test this hypothesis, we isolated and cultured renal PTCs from 22-mo-old F344 rats and their controls, normotensive 4-mo-old F344 rats, and SHRs and their controls, normotensive Wistar-Kyoto (WKY) rats. The results demonstrate that in both hypertensive models (SHR and aged F344), NHERF-1 expression, dopamine-mediated phosphorylation of NKA, and ouabain-inhibitable K+ transport are reduced. Transfection of NHERF-1 into PTCs from aged F344 and SHRs restored dopamine-mediated inhibition of NKA. These results suggest that decreased renal NHERF-1 expression contributes to the impaired dopamine-mediated inhibition of NKA in PTCs from animal models of hypertension.

Low dose ouabain stimulates NaK ATPase α1 subunit association with angiotensin II type 1 receptor in renal proximal tubule cells.

Our laboratory has recently demonstrated that low concentrations of ouabain increase blood pressure in rats associated with stimulation of NaK ATPase activity and activation of the Src signaling cascade in NHE1-dependent manner. Proteomic analysis of human kidney proximal tubule cells (HKC11) suggested that the Angiotensin II type 1 receptor (AT1R) as an ouabain-associating protein. We hypothesize that ouabain-induced stimulation of NaK ATPase activity is mediated through AT1R. To test this hypothesis, we examined the effect of ouabain on renal cell angiotensin II production, the effect of AT1R inhibition on ouabain-stimulated NKA activity, and the effect of ouabain on NKA-AT1R association. Ouabain increased plasma angiotensin II levels in rats treated with ouabain (1µg/kg body wt./day) for 9days and increased angiotensin II levels in cell culture media after 24h treatment with ouabain in human (HKC11), mouse (MRPT), and human adrenal cells. Ouabain 10pM stimulated NKA-mediated 86Rb uptake and phosphorylation of EGFR, Src, and ERK1/2. These effects were prevented by the AT1R receptor blocker candesartan. FRET and TIRF microscopy using Bodipy-labeled ouabain and mCherry-NKA or mCherry-AT1R demonstrated association of ouabain with AT1R and NKA. Further our FRET and TIRF studies demonstrated increased association between AT1R and NKA upon treatment with low dose ouabain. We conclude that ouabain stimulates NKA in renal proximal tubule cells through an angiotensin/AT1R-dependent mechanism and that this pathway contributes to cardiac glycoside associated hypertension.

Role of Na+/H+ exchanger regulatory factor 1 in forward trafficking of the type IIa Na+-Pi cotransporter.

Na+/H+ exchanger regulatory factor (NHERF1) plays a critical role in the renal transport of phosphate by binding to Na+-Pi cotransporter (NpT2a) in the proximal tubule. While the association between NpT2a and NHERF1 in the apical membrane is known, the role of NHERF1 to regulate the trafficking of NpT2a has not been studied. To address this question, we performed cell fractionation by sucrose gradient centrifugation in opossum kidney (OK) cells placed in low-Pi medium to stimulate forward trafficking of NpT2a. Immunoblot analysis demonstrated expression of NpT2a and NHERF1 in the endoplasmic reticulum (ER)/Golgi. Coimmunoprecipitation demonstrated a NpT2a-NHERF1 interaction in the ER/Golgi. Low-Pi medium for 4 and 8 h triggered a decrease in NHERF1 in the plasma membrane with a corresponding increase in the ER/Golgi. Time-lapse total internal reflection fluorescence imaging of OK cells placed in low-Pi medium, paired with particle tracking and mean square displacement analysis, indicated active directed movement of NHERF1 at early and late time points, whereas NpT2a showed active movement only at later times. Silence of NHERF1 in OK cells expressing green fluorescent protein (GFP)-NpT2a resulted in an intracellular accumulation of GFP-NpT2a. Transfection with GFP-labeled COOH-terminal (TRL) PDZ-binding motif deleted or wild-type NpT2a in OK cells followed by cell fractionation and immunoprecipitation confirmed that the interaction between NpT2a and NHERF1 was dependent on the TRL motif of NpT2a. We conclude that appropriate trafficking of NpT2a to the plasma membrane is dependent on the initial association between NpT2a and NHERF1 through the COOH-terminal TRL motif of NpT2a in the ER/Golgi and requires redistribution of NHERF1 to the ER/Golgi.