Mitochondrial uncoupling proteins protect human airway epithelial ciliated cells from oxidative damage.

Apical cilia on epithelial cells defend the lung by propelling pathogens and particulates out of the respiratory airways. Ciliated cells produce ATP that powers cilia beating by densely grouping mitochondria just beneath the apical membrane. However, this efficient localization comes at a cost because electrons leaked during oxidative phosphorylation react with molecular oxygen to form superoxide, and thus, the cluster of mitochondria creates a hotspot for oxidant production. The relatively high oxygen concentration overlying airway epithelia further intensifies the risk of generating superoxide. Thus, airway ciliated cells face a unique challenge of producing harmful levels of oxidants. However, surprisingly, highly ciliated epithelia produce less reactive oxygen species (ROS) than epithelia with few ciliated cells. Compared to other airway cell types, ciliated cells express high levels of mitochondrial uncoupling proteins, UCP2 and UCP5. These proteins decrease mitochondrial protonmotive force and thereby reduce production of ROS. As a result, lipid peroxidation, a marker of oxidant injury, decreases. However, mitochondrial uncoupling proteins exact a price for decreasing oxidant production; they decrease the fraction of mitochondrial respiration that generates ATP. These findings indicate that ciliated cells sacrifice mitochondrial efficiency in exchange for safety from damaging oxidation. Employing uncoupling proteins to prevent oxidant production, instead of relying solely on antioxidants to decrease postproduction oxidant levels, may offer an advantage for targeting a local area of intense ROS generation.

WNK Inhibition Increases Surface Liquid pH and Host Defense in Cystic Fibrosis Airway Epithelia.

In cystic fibrosis (CF), reduced HCO3- secretion acidifies the airway surface liquid (ASL), and the acidic pH disrupts host defenses. Thus, understanding the control of ASL pH (pHASL) in CF may help identify novel targets and facilitate therapeutic development. In diverse epithelia, the WNK (with-no-lysine [K]) kinases coordinate HCO3- and Cl- transport, but their functions in airway epithelia are poorly understood. Here, we tested the hypothesis that WNK kinases regulate CF pHASL. In primary cultures of differentiated human airway epithelia, inhibiting WNK kinases acutely increased both CF and non-CF pHASL. This response was HCO3- dependent and involved downstream SPAK/OSR1 (Ste20/SPS1-related proline-alanine-rich protein kinase/oxidative stress responsive 1 kinase). Importantly, WNK inhibition enhanced key host defenses otherwise impaired in CF. Human airway epithelia expressed two WNK isoforms in secretory cells and ionocytes, and knockdown of either WNK1 or WNK2 increased CF pHASL. WNK inhibition decreased Cl- secretion and the response to bumetanide, an NKCC1 (sodium-potassium-chloride cotransporter 1) inhibitor. Surprisingly, bumetanide alone or basolateral Cl- substitution also alkalinized CF pHASL. These data suggest that WNK kinases influence the balance between transepithelial Cl- versus HCO3- secretion. Moreover, reducing basolateral Cl- entry may increase HCO3- secretion and raise pHASL, thereby improving CF host defenses.

Cellular and molecular architecture of submucosal glands in wild-type and cystic fibrosis pigs.

Submucosal glands (SMGs) protect lungs but can also contribute to disease. For example, in cystic fibrosis (CF), SMGs produce abnormal mucus that disrupts mucociliary transport. CF is an ion transport disease, yet knowledge of the ion transporters expressed by SMG acini, which produce mucus, and SMG ducts that carry it to the airway lumen is limited. Therefore, we isolated SMGs from newborn pigs and used single-cell messenger RNA sequencing, immunohistochemistry, and in situ hybridization to identify cell types, gene expression, and spatial distribution. Cell types and transcript levels were the same in non-CF and CF SMGs, suggesting that loss of epithelial anion secretion rather than an intrinsic cell defect causes CF mucus abnormalities. Gene signatures of acinar mucous and acinar serous cells revealed specialized functions in producing mucins and antimicrobials, respectively. However, surprisingly, these two cell types expressed the same ion transporters and neurohumoral receptors, suggesting the importance of balancing mucin and liquid secretion to produce optimal mucus properties. SMG duct cell transcripts suggest that they secrete HCO3- and Cl-, and thus have some similarity to pancreatic ducts that are also defective in CF. These and additional findings suggest the functions of the SMG acinus and duct and provide a baseline for understanding how environmental and genetic challenges impact their contribution to lung disease.

Initial report of the genitourinary and gastrointestinal toxicity of post-prostatectomy proton therapy for prostate cancer patients undergoing adjuvant or salvage radiotherapy.

PURPOSE: To report acute and late genitourinary (GU) and gastrointestinal (GI) toxicities associated with post-prostatectomy proton therapy (PT). METHODS: The first 100 consecutive patients from 2010 to 2016 were retrospectively assessed. Baseline characteristics, prospectively graded CTCAE v4.0 toxicities, and patient-reported outcomes were reported. Late outcomes were reported for 79 patients with 3 months minimum follow up. Toxicity-free survival Kaplan-Meier curves were estimated. Logistic regression assessed associations between toxicities and clinical and treatment characteristics (p < .05 significance). RESULTS: Median age, months after surgery, and months of follow-up were respectively 64 years (range 42-77), 25 (5-216), and 25 (0-47). PT received was 70.2 Gy (RBE) (89%), salvage (93%), prostate bed only (80%), pencil beam scanning (86%), with IMRT (31%), and with androgen deprivation (34%). Acute and late maximum toxicities, respectively were: GU grade 0 (14%; 18%), 1 (71%; 62%), 2 (15%; 20%), ≥3 (0), and GI: grade 0 (66%; 73%), 1 (34%; 27%), ≥2 (0). Toxicity-free survival at 24 months was GU grade 2 (83%) and GI grade 1 (74%). Mean (±std dev) baseline International Prostate Symptom Score (IPSS), International Index of Erectile Function, and Expanded Prostate Cancer Index Composite bowel function and bother were 6.6 ± 6.1, 10.5 ± 7.3, 90.9 ± 10.8, 93.3 ± 11.2, respectively, and largely unchanged at 2 years: 6.3 ± 3.6, 11.1 ± 6.3, 92.8 ± 5.8, and 90.9 ± 10.3. On multivariate analysis, baseline IPSS (p = .009) associated with GU grade 2 acute toxicity. Bladderless-CTV median dose, V30, and V40 associated with GU grade 2 acute toxicity and maximum dose with late (Ps <0.05). For GI, on multivariate analysis, baseline bowel function (p = .033) associated with acute grade 1 toxicity. Rectal minimum and median dose, V10, and V20, and anterior rectal wall median dose and V10 through V65 associated with acute grade 1 GI toxicity (Ps < .05). CONCLUSIONS: Post-prostatectomy PT for prostate cancer is feasible with a favorable GU and GI toxicity profile acutely and through early follow up.

Sociodemographic disparities in the utilization of proton therapy for prostate cancer at an urban academic center.

PURPOSE: Despite increasing use, proton therapy (PT) remains a relatively limited resource. The purpose of this study was to assess clinical and demographic differences in PT use for prostate cancer compared to intensity modulated radiation therapy (IMRT) at a single institution. METHODS AND MATERIALS: All patients with low- and intermediate-risk prostate cancer (N = 633) who underwent definitive radiation therapy between 2010 and 2015 were divided into PT (n = 508) and IMRT (n = 125) comparison groups and compared using χ2 and independent sample t tests. Univariable and multivariable logistic regression analyses were conducted to assess the associations between PT use and demographic, clinical, and treatment characteristics. RESULTS: The PT and IMRT cohorts varied by age, race, poverty, distance, treatment year, and treating physician. Patients who underwent IMRT were more likely to be older (mean age, 66 vs. 68 years), black (51% vs. 75%), and living in poverty or close to the facility (mean distance between residence and facility, 90 vs. 21 miles; P < .05). Prostate-specific antigen, prostate volume, and International Index of Erectile Function were significantly higher in the IMRT cohort (P < .05), but insurance type, risk group, tumor stage, Gleason score, and patient-reported urinary and bowel scores did not differ significantly (P > .05). Patients who underwent PT were more likely to receive hypofractionated therapy and less likely to receive androgen deprivation therapy (P < .01). On multivariable analysis, black (odds ratio [OR], 0.29; 95% confidence interval [CI], 0.15-0.57) and other race (OR, 0.42; 95% CI, 0.20-0.90); distance (OR, 1.14; 95% CI, 1.06-1.24); treatment years 2011 (OR, 4.87; 95% CI, 2.23-10.6), 2012 (OR, 8.27; 95% CI, 3.43-19.9), and 2014 (OR, 4.44; 95% CI, 1.94-10.2) relative to 2010; and a single treating physician (OR, 0.38; 95% CI, 0.18-0.81) relative to the reference physician with the highest rate of use were associated with PT use, whereas clinical factors such as prostate-specific antigen, prostate volume, International Index of Erectile Function, and androgen deprivation therapy were not. CONCLUSION: Sociodemographic disparities exist in PT use for prostate cancer at an urban academic institution. Further investigation of potential barriers to access is warranted to ensure equitable distribution across all demographic groups.

CD19-negative B-lineage acute lymphoblastic leukemia: A diagnostic and therapeutic challenge.

B-lineage acute lymphoblastic leukemia (B-ALL) is an aggressive neoplasm of B-lymphocyte precursors that express the pan B-cell marker CD19 in all the cases. Rarely, a case may be assigned as B-lineage even if CD19 is negative. Here, a 16-year-old male presented with complaints of pain abdomen, on and off fever, joint pain, and hepatosplenomegaly for 2 months. Bone marrow examination was suggestive of acute leukemia with numerous leukoblasts on aspiration. On flow cytometry, gated blast population was negative for CD19, cytoCD3, and myeloperoxidase MPO and positive for CD34, TdT, HLA-DR, CD22, CD79a, and CD10. Immunohistochemistry study showed positivity for TdT, CD34, CD10 (focal), and PAX 5 and negativity for CD20, CD3, MPO, CD117, and CD68. Lack of awareness of negative CD19 expression in B-ALL can lead to incorrect immunophenotypic diagnosis, treatment, and monitoring of B-ALL. Proper diagnosis should be based on clinical features, immunophenotypic profiles, immunohistochemistry findings, and molecular analysis.

Modulation of nutritional and antioxidant potential of seeds and pericarp of pea pods treated with microbial consortium.

Microbial populations have diverse roles within rhizosphere where interactions among distinct microorganisms along with the host may lead to mutualistic associations. The present study aimed to investigate the nutritional and antioxidant qualities of seeds and pericarp of pea raised from seeds treated with beneficial microbes namely Bacillus subtilis BHHU100, Trichoderma harzianum TNHU27 and Pseudomonas aeruginosa PJHU15 either singly and/or in consortia. A significant increase in total phenolic, flavonoid, ascorbic acid and protein contents, free radical scavenging activity, hydroxyl radical scavenging activity, iron chelation and reducing power were observed in the seeds and pericarp of pods treated with a consortium of microbes in comparison to control pods. Also, the differential accumulation of phenolic compounds, namely, shikimic acid, gallic acid, tannic acid, syringic acid, p-coumaric acid, quercetin and kaempferol, was observed from the HPLC chromatogram of the seed extracts of different treatments. We especially emphasized on dietary importance of the pod pericarp, other than seeds, along with their modulation by microbial consortium. The study also highlights the role of beneficial microbes in improving nutritional value by providing protection against oxidative stress.