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Hematopoietic cell transplant for sickle cell disease is curative but is associated with life threatening complications most of which occur within the first 2 years after transplantation. In the current era with interest in gene therapy and gene editing we felt it timely to report on sickle cell disease transplant recipients who were alive for at least 2-year after transplantation, not previously reported. Our objectives were to (1) report the conditional survival rates of patients who were alive for 2 or more years after transplantation (2) identify risk factors for death beyond 2 years after transplantation and (3) compare all-cause mortality risks to those of an age-, sex- and race-matched general population in the United States. By limiting to 2-year survivors, we exclude deaths that occur as a direct consequence of the transplantation procedure. De-identified records of 1149 patients were reviewed from a publicly available data source and 950 patients were eligible (https://picsure.biodatacatalyst.nhlbi.nih.gov). All analyses were performed in this secure cloud environment using the available statistical software package(s). The validity of the public database was confirmed by reproducing results from an earlier publication. Conditional survival estimates were obtained using the Kaplan-Meier method for the sub-cohort that had survived a given length (x) of time after transplantation. Cox regression models were built to identify risk factors associated with mortality beyond 2 years after transplantation. The standardized relative mortality risk (SMR) or the ratio of observed to expected number of deaths, was used to quantify all-cause mortality risk after transplantation and compared to age, race and sex-matched general population. Person-years at risk were calculated from an anchor date (i.e., 2-, 5- and 7-years) after transplantation until date of death or last date known alive. The expected number of deaths was calculated using age, race and sex-specific US mortality rates. The median follow up was 5 years (range 2-20) and 300 (32%) patients were observed for more than 7 years. Among those who lived for at least 7 years after transplantation the 12-year probability of survival was 97% (95% CI, 92%-99%). Compared to an age-, race- and sex-matched US population, the risk for late death after transplantation was higher as late as 7 years after transplantation (hazard ratio (HR) 3.2; P= .020) but the risk receded over time. Risk factors for late death included age at transplant and donor type. For every 10-year increment in patient age, an older patient was 1.75 times more likely to die than a younger patient (P= .0004). Compared to HLA-matched siblings the use of other donors was associated with higher risk for late death (HR 3.49; P= .003). Graft failure (beyond 2-years after transplantation) was 7% (95% CI, 5%-9%) and graft failure was higher after transplantation of grafts from donors who were not HLA-matched siblings (HR 2.59, P< .0001). Long-term survival after transplantation is excellent and support this treatment as a cure for sickle cell disease. The expected risk for death recedes over time but the risk for late death is not negligible., (Copyright © 2022 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

Clarke and Beck argue that the approximate number system (ANS) represents rational numbers, like 1/3 or 3.5. I think this claim is not supported by the evidence. Rather, I argue, ANS should be interpreted as representing natural numbers and ratios among them; and we should view the contents of these representations are genuinely approximate.

Objectives: To compare registry and electronic health record (EHR) data mining approaches for cohort ascertainment in patients with pediatric pulmonary hypertension (PH) in an effort to overcome some of the limitations of registry enrollment alone in identifying patients with particular disease phenotypes., Study Design: This study was a single-center retrospective analysis of EHR and registry data at Boston Children's Hospital. The local Informatics for Integrating Biology and the Bedside (i2b2) data warehouse was queried for billing codes, prescriptions, and narrative data related to pediatric PH. Computable phenotype algorithms were developed by fitting penalized logistic regression models to a physician-annotated training set. Algorithms were applied to a candidate patient cohort, and performance was evaluated using a separate set of 136 records and 179 registry patients. We compared clinical and demographic characteristics of patients identified by computable phenotype and the registry., Results: The computable phenotype had an area under the receiver operating characteristics curve of 90% (95% CI, 85%-95%), a positive predictive value of 85% (95% CI, 77%-93%), and identified 413 patients (an additional 231%) with pediatric PH who were not enrolled in the registry. Patients identified by the computable phenotype were clinically distinct from registry patients, with a greater prevalence of diagnoses related to perinatal distress and left heart disease., Conclusions: Mining of EHRs using computable phenotypes identified a large cohort of patients not recruited using a classic registry. Fusion of EHR and registry data can improve cohort ascertainment for the study of rare diseases., Trial Registration: ClinicalTrials.gov: NCT02249923., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Introduction: Despite growing interest in integrating patient-reported outcome (PRO) measures of symptoms and functional status into routine cancer care, little attention has been paid to patients' and clinicians' perceptions of acceptability and value., Methods: A two-phase qualitative study was conducted to develop a web-based PRO screening system with 21 items assessing symptoms (e.g., nausea) and functional status. Phase 1 involved cognitive interviews with 35 cancer outpatients (n=9 breast chemotherapy, radiation for prostate (n=8) or head and neck cancer (n=10), and n=8 bone marrow transplant [BMT]). In Phase 2, we evaluated the acceptability and perceived value of reviewing a PRO measure during real-time clinical encounters with 39 additional outpatients (n=10 breast, n=9 head and neck, n=10 prostate, n=10 BMT) and 12 clinicians (n=3 breast, n=2 head and neck, n=4 prostate, n=3 BMT). At least 20% of patients were ≥60 years, African American, or ≤ high school., Results: Patients felt that their PRO summary of symptoms and functional status was helpful in discussing health issues with clinicians (92%), wanted to review their results with clinicians during future visits (82%), and would recommend it to other patients (87%). Clinicians found the PRO summary to be easy to interpret (83%), most helpful for documenting the Review of Symptoms (92%), and would recommend it to future patients (92%). Over 90% of clinicians reported that consultation time did not increase., Conclusion: Both cancer patients and clinicians reported that discussing a PRO summary of symptoms and functional status during an outpatient visit was useful, acceptable, and feasible.

Background: Understanding the impact of the cancer care system on racial/ethnic disparities in health-related quality of life (HRQOL) is increasingly important as the number of cancer survivors in the United States grows. The authors prospectively assessed changes in HRQOL before and after a first cancer diagnosis among non-Hispanic whites (NHWs), African Americans (AAs), Hispanics, and Asians in a cohort of Medicare beneficiaries with and without cancer., Methods: Data from the Surveillance, Epidemiology, and End Results linked with the Medicare Health Outcomes Survey were used to identify 1778 individuals aged ≥65 years with prostate, breast, or colorectal cancer from 1998 to 2007. The Medical Outcomes Trust Short Form 36 (SF-36) instrument was used to measure HRQOL. By using propensity scores, each patient with cancer (case) was matched to 5 individuals without cancer (noncancer controls), and differences in HRQOL according to race/ethnicity were assessed. Mixed effects analysis of covariance models was used to assess differences in HRQOL, adjusting for baseline HRQOL, demographics, and self-reported comorbid conditions while controlling for each individual's managed care plan. Stratified analyses were used to assess racial/ethnic disparities between cases and noncancer controls., Results: Before cancer diagnosis, NHWs had better HRQOL scores than AAs and Hispanics on the Role-Physical and Role-Emotional SF-36 subscales. Cancer diagnosis/treatment negatively impacted individuals' lives regardless of race/ethnicity. However, among cases, gaps between racial/ethnic groups narrowed (compared with controls) before and after cancer diagnosis for some SF-36 HRQOL measures., Conclusions: Racial/ethnic HRQOL gaps exist among cancer survivors but may narrow because of exposure to the cancer care system. Further research to understand why this occurs will help inform initiatives to manage the impact of cancer on HRQOL among elderly cancer survivors., (© 2014 American Cancer Society.)

Background: Data on health-related quality of life (HRQoL) changes among Americans aged ≥65 following colorectal cancer (CRC) diagnosis and treatment are limited. This study compared HRQoL changes among CRC patients across stages from before to after diagnosis with matched noncancer controls., Methods: This population-based study used the Surveillance, Epidemiology, and End Results Medicare Health Outcomes Survey (MHOS) data set (1998-2007). Medicare Advantage beneficiaries diagnosed with CRC between their baseline and follow-up MHOS (n = 349) were matched to noncancer controls (n = 1745) using propensity scores. Mixed-effects analysis of covariance models estimated changes in HRQoL (measured by the Medical Outcomes Study Short Form-36/Veterans RAND 12-item Survey) and the ability to perform 6 activities of daily living (ADLs) between baseline and follow-up. Logistic regression models estimated odds ratios for ADL impairments and major depressive disorder (MDD) risk., Results: Mean time between CRC diagnosis and follow-up MHOS was 12.3 ± 9.8 months. Compared with controls, CRC patients had significantly lower scores in all physical and mental health domains at follow-up. The greatest decrements were observed in physical health and were largely driven by declines in the 6 months postdiagnosis and in stage III and IV patients. At follow-up, CRC patients had greater overall ADL impairment and difficulty with dressing, eating, and getting in/out of chairs. CRC patients, particularly stage IV patients, had greater odds of being at risk for MDD relative to controls., Conclusions: This study further underscores the adverse effects of CRC on physical health and the need to support older Americans' basic self-care needs, with attention to later-stage patients' increased debility., (© 2014 American Cancer Society.)

Objective: To improve and individualise estimates of treatment outcomes for men diagnosed with prostate cancer, we examined the impact of baseline comorbidity on health-related quality of life (HRQL) outcomes in an analysis of two pooled, prospective cohort studies., Patients and Methods: We studied 697 patients from three academic hospitals who received radical prostatectomy (RP), external beam radiation therapy (EBRT), or brachytherapy (BT). Measures of patient-reported bowel, urinary, and sexual symptoms along with physical and mental health were prospectively collected before treatment and 3, 12, 24, and 36 months after treatment. We assessed baseline comorbidity by the validated Index of Co-Existent Disease (ICED), abstracted from medical records. Regression mixed-models were built for each treatment group and HRQL outcome controlling for baseline age, education, marital status, risk group and patient-reported general health., Results: About 71% of patients had one or more comorbid conditions at baseline. After adjusting for covariates, we found baseline comorbidity was independently associated with poorer sexual function after BT (P = 0.04) and RP (P = 0.03) but not EBRT (P = 0.35). Physical health was significantly worse for men receiving BT with more comorbidities (P = 0.02). Baseline comorbid conditions were not associated with urinary incontinence or bowel functioning., Conclusions: Comorbidity at baseline is significantly associated with poorer sexual function after prostate BT or RP. This information may help patients and their physicians anticipate outcomes after surgical and radiation treatments., (© 2014 The Authors. BJU International © 2014 BJU International.)

Background: Health care providers have little population-based evidence about health-related quality of life (HRQOL) changes, from the pre- to postdiagnosis period, and treatment-related recovery time for women aged 65 years and older diagnosed with breast cancer., Methods: Older women with and without breast cancer completed self-reports of HRQOL at baseline and 2 years later as part of annual Medicare Health Outcomes Surveys (MHOS). MHOS was linked to Surveillance, Epidemiology, and End Results registries, which were used to categorize women with breast cancer by treatment type (breast-conserving surgery, breast-conserving surgery plus radiation, mastectomy) and time since diagnosis at follow-up. Each cancer case diagnosed in 1998 through 2007 (N = 542) was matched to 5 women without cancer (N = 2710) using propensity score matching. Analysis of covariance models examined changes in HRQOL, adjusting for demographics and initial functioning., Results: Older women within 6 months of diagnosis had greater declines than women without cancer in SF-36 Physical (-5.8 vs -1.8) and Mental (-3.6 vs -0.7) Component Summary scores, General Health (-12.3 vs -4.6), Vitality (-11.0 vs -2.2), Bodily Pain (-8.5 vs -2.1), Social Functioning (-15.1 vs -3.3), Role-Physical (-26.5 vs -3.9), and Role-Emotional (-13.1 vs -3.1) scores (all P < .05). By approximately 1 year, women with and without breast cancer had similar HRQOL. Comparable declines in Physical Component Summary and Role-Physical occurred across treatment types., Conclusions: Women aged 65 years and older diagnosed with breast cancer should be counseled that survivors within 6 months of diagnosis are vulnerable to HRQOL declines, compared to women without breast cancer, but that decrements generally wane after 12 months., (© 2014 American Cancer Society.)

Purpose: An essential aspect of patient-centered outcomes research (PCOR) and comparative effectiveness research (CER) is the integration of patient perspectives and experiences with clinical data to evaluate interventions. Thus, PCOR and CER require capturing patient-reported outcome (PRO) data appropriately to inform research, healthcare delivery, and policy. This initiative's goal was to identify minimum standards for the design and selection of a PRO measure for use in PCOR and CER., Methods: We performed a literature review to find existing guidelines for the selection of PRO measures. We also conducted an online survey of the International Society for Quality of Life Research (ISOQOL) membership to solicit input on PRO standards. A standard was designated as "recommended" when >50 % respondents endorsed it as "required as a minimum standard.", Results: The literature review identified 387 articles. Survey response rate was 120 of 506 ISOQOL members. The respondents had an average of 15 years experience in PRO research, and 89 % felt competent or very competent providing feedback. Final recommendations for PRO measure standards included: documentation of the conceptual and measurement model; evidence for reliability, validity (content validity, construct validity, responsiveness); interpretability of scores; quality translation, and acceptable patient and investigator burden., Conclusion: The development of these minimum measurement standards is intended to promote the appropriate use of PRO measures to inform PCOR and CER, which in turn can improve the effectiveness and efficiency of healthcare delivery. A next step is to expand these minimum standards to identify best practices for selecting decision-relevant PRO measures.

Purpose: Children with cancer experience significant toxicities while undergoing treatment. Documentation of adverse events (AEs) in clinical trials is mandated by federal agencies. Although many AEs are subjective, the current standard is clinician reporting. Our long-term goal is to create and validate a self-report measure of subjective AEs for children aged 7 years and older that will inform AE reporting for the National Cancer Institute's Common Terminology Criteria for Adverse Events (CTCAE). This content validation study aimed to identify which of the AEs in the current CTCAE should be included in a pediatric self-report measure., Methods: We sought expert panel review and consensus among 187 pediatric clinicians from seven Children's Oncology Group institutions to determine which of the 790 AEs are amenable to child self-report. Two survey iterations were used to identify suitable AEs, and clinician agreement estimated by the content-validity ratio (CVR) was assessed., Results: Response rates for surveys 1 and 2 were 72% and 67%, respectively. After the surveys, 64 CTCAE terms met the criteria of being subjective, relevant for use in pediatric cancer trials, and amenable to self-report by a child. The most frequent reasons for removal of CTCAE terms were that they relied on laboratory or clinical measures or were not applicable to children., Conclusion: The 64 CTCAE terms will be translated into child-friendly terms as the basis of the child-report toxicity measure. Ultimately, systematic collection of these data will improve care by enhancing the accuracy and completeness of treatment toxicity reports for childhood cancer., (Copyright © 2013 Wiley Periodicals, Inc.)

The effect of environmental acidity on the induction of apoptosis by heat was investigated. Human colorectal tumour RKO.C cells, carrying wild-type p53 and isogenic RC10.1 cells deficient in p53, were heated at 42.0 degrees C for 1 h in pH 7.5 or pH 6.6 medium and the apoptosis was assessed based on the flow cytometic determination of DNA content, DNA fragmentation, and PARP cleavage. The degree of apoptosis after heating in pH6.6 medium was greater than that in pH 7.5 medium in both RKO.C cells and RC10.1 cells. When heated in the same pH medium, more apoptosis occurred in the RC10.1 cells than in the RKO.C cells. Heating increased the expression of p53 protein and p21 protein markedly in RKO.C cells and slightly in RC10.1 cells. Expression of these proteins was slightly greater in pH 7.5 medium than in pH 6.6 medium. The expressions of Bax protein and Bcl-2 protein, which are known to control apoptosis, were not altered by heating. It was concluded that an acidic environment enhances heat-induced apoptosis. It was also concluded that heat-induced apoptosis is lessened by p53 and that Bcl-2 and Bax are not involved in the induction of apoptosis by hyperthermia.

Arsenic trioxide, As2O3 (ATO), has been found to be an effective chemotherapeutic for acute promyelocytic leukemia but its effect on solid tumors has not been fully explored. In the present report, we describe our observation that ATO is a potent antivascular agent and that it markedly enhances the effect of hyperthermia on tumors. The tumor blood perfusion in SCK tumors of A/J mice and FSaII tumors of C3H mice was significantly suppressed for up to 24 hours after an i.p. injection of 8 mg/kg ATO. ATO was also found to be able to increase the thermosensitivity of tumor cells in vitro. As a probable consequence of these effects, ATO treatment markedly increased the tumor growth delay caused by hyperthermia at 41.5 to 42.5 degrees C. Immunohistochemical staining of tumor tissue revealed that the expression levels of several adhesion molecules and TNFalpha are noticeably increased in tumors 2 to 6 hours after systemic ATO treatment. It is concluded that ATO is potentially useful to enhance the effect of hyperthermia on tumors at a clinically relevant temperature.

The successful use and placement of subgingival resin-ionomer restorations in both anterior root and molar furcation defects are demonstrated in this article. Sustained tissue health and minimal probing depths at the surgical site demonstrate clinical success. These case reports illustrate the continued success of alternative treatment procedures for restoring subgingival mechanical root or periodontal lesions.

We investigated the effect of an acidic environment on the radiation-induced G2/M arrest and apoptosis using RKO.C human colorectal cancer cells expressing wild-type p53 and RC10.1 cells, a subline of RKO.C cells deficient in p53 as well as p53+/+ MEFs and p53-/- MEFs (mouse embryonic fibroblasts). The cells were irradiated with 4 Gy or 12 Gy of gamma-rays in pH 7.5 medium or pH 6.6 medium. p53 accentuated the progression of cells from radiation-induced G2/M arrest to apoptosis and the pH 6.6 environment suppressed the progression of cells through G2/M-phase to apoptosis after irradiation. Further analysis indicated that the radiation-induced G2/M arrest was due mainly to G2 arrest in both pH 7.5 and pH 6.6. Therefore, it was concluded that p53 enhances, and an acidic environment suppresses, the exit of cells from radiation-induced G2 arrest by altering cyclin B1-Cdc2 kinase activity.

Apoptosis and cell cycle progression in HL60 cells irradiated in an acidic environment were investigated. Apoptosis was determined by TUNEL staining, PARP cleavage, DNA fragmentation, and flow cytometry. The majority of the apoptosis that occurred in HL60 cells after 4 Gy irradiation took place after G(2)/M-phase arrest. When irradiated with 12 Gy, a fraction of the cells underwent apoptosis in G(1) and S phases while the rest of the cells underwent apoptosis in G(2)/M phase. The apoptosis caused by 4 and 12 Gy irradiation was transiently suppressed in medium at pH 7.1 or lower. An acidic environment was found to perturb progression of irradiated cells through the cell cycle, including progression through G(2)/ M phase. Thus it was concluded that the suppression of apoptosis in the cells after 4-12 Gy irradiation in acidic medium was due at least in part to a delay in cell cycle progression, particularly the prolongation of G(2)/M-phase arrest. Irradiation with 20 Gy indiscriminately caused apoptosis in all cell cycle phases, i.e. G(1), S and G(2)/M phases, rapidly in neutral pH medium and relatively slowly in acidic pH medium. The delay in apoptosis in acidic medium after 20 Gy irradiation appeared to result from mechanisms other than prolonged G(2)/ M-phase arrest.

An exposure of HL-60 human promyelocytic leukaemia cells to acidic media with pH 6.2-6.6 caused an up-regulation of Bax protein expression within 2 h, which lasted for longer than 6 h. On the other hand, the apoptosis, as judged from PARP cleavage, DNA fragmentation and flow cytometric determination of cell population with sub-G1 DNA content, occurred after the cells were incubated in the acidic media for longer than 4 h. The PARP cleavage and DNA fragmentation in the cells exposed to an acidic environment could be effectively suppressed by inhibitors specific for ICE or CPP32, indicating that activation of these caspases is an essential step in acidic stress-induced apoptosis. It has been known that Bax is involved in the activation of caspases. Taken together, it appears that acidic stress first up-regulates Bax protein thereby activating caspases followed by PARP cleavage and DNA fragmentation. The observation that inhibition of either ICE or CPP32 could suppress acidic stress-induced apoptosis suggested that ICE activates pro-CPP32, which then cleaves PARP. Flow cytometric analysis indicated that acidic stress-induced apoptosis occurs mainly in G1 cells. The finding in the present study demonstrated that acidic intra-tumour environment may markedly perturb the tumour cell proliferation and tumour growth.

The effects of an acidic environment on the induction of apoptosis by 42 degrees C hyperthermia were investigated. An acidic environment (pH 6.6) enhanced the hyperthermia-induced apoptosis in HL-60 human promyelocytic leukemia cells as judged by the DNA fragmentation, flow cytometric analysis of DNA content, and cleavage of poly(ADP-ribose) polymerase. Hyperthermia exerted no effect on the expression of Bcl-2 and Bax, regardless of the environmental acidity during heating. The time of increase in apoptosis after heating coincided with the time of decrease in the G1-phase cell population. It seemed that the increase in heat-induced apoptosis in HL-60 cells in an acidic environment was due to a direct increase in the proteolytic cleavage of poly(ADP-ribose) polymerase by acidic caspases without the involvement of Bcl-2 and Bax, and that heat-induced apoptosis occurred during G1 phase in HL-60 cells.

Malignant gliomas have been associated with a high rate of glycolytic activity which is believed necessary to sustain cellular function and integrity. Since lonidamine (LND) is believed to reduce tumor glucose utilization by inhibition of the mitochondrially-bound glycolytic enzyme hexokinase (HK), 31P magnetic resonance spectroscopy (MRS) was used to noninvasively follow the effects of LND on both tumor pH and the high-energy phosphate metabolites: ATP, phosphocreatine (PCr) and inorganic phosphate (Pi) in subcutaneous rat 9L gliosarcomas. 31P tumor spectra acquired in 5 min intervals pre- and post LND administration of 50 and 100 mg/kg, i.p. revealed an acidotic pH shift of -0.25 and -0.45 pH units, respectively within 30 min post administration. The ATP/Pi ratio of 9L tumors decreased to 40% of control and Pi levels increased to 280% of control over a 3 hr period. LND exerted no effect on tumor blood flow and mean arterial blood pressure. Brain and muscle metabolite levels and pH were also unaffected by LND. In vitro measurements of cultured 9L tumor cell intra- and extracellular lactate, pentose phosphate pathway (PPP) and hexokinase (HK) activities suggest that the mode of action of LND involves inhibition of lactate efflux and intracellular acidification. The selective reduction of tumor energy metabolites and pH by LND may be exploitable for sensitizing gliomas to radiation, chemotherapy or hyperthermia.

Purpose: The effect of environmental pH on the radiation-induced apoptosis in tumor cells in vitro was investigated., Methods and Materials: Mammary adenocarcinoma cells of A/J mice (SCK cells) were irradiated with gamma-rays using a 137Cs irradiator and incubated in media of different pHs. After incubation at 37 degrees C for 24-120 h the extent of apoptosis was determined using agarose gel electrophoresis, TdT-mediated dUTP-biotin nick end labeling (TUNEL) staining, flow cytometry, and release of 3H from 3H-thymidine labeled cells. The clonogenicity of the cells irradiated in different pH medium was determined, and the progression of cells through the cell cycle after irradiation in different pHs was also determined with flow cytometry., Results: Irradiation with 2-12 Gy of gamma-rays induced apoptosis in SCK cells in pH 7.5 medium within 48 h as judged from the results of four different assays mentioned. Radiation-induced apoptosis declined as the medium pH was lowered from 7.5 to 6.4. Specifically, the radiation-induced degradation of DNA including the early DNA breaks, as determined with the TUNEL method, progressively declined as the medium pH was lowered so that little DNA fragmentation occurred 48 h after irradiation with 12 Gy in pH 6.6 medium. When the cells were irradiated and incubated for 48 h in pH 6.6 medium and the medium was replaced with pH 7.5 medium, DNA fragmentation promptly occurred. DNA fragmentation also occurred even in pH 6.6 medium when the cells were irradiated and maintained in pH 7.5 medium for 8 h or longer post-irradiation before incubation in pH 6.6 medium. The radiation-induced G2 arrest in pH 6.6 medium lasted markedly longer than that in pH 7.5 medium., Conclusion: Radiation-induced apoptosis in SCK cells in vitro is reversibly suppressed in an acidic environment. Taking the results of four different assays together, it was concluded that early step(s) in the apoptotic pathway, probably the DNA break or upstream of DNA break, is reversibly halted by an acidic environment in irradiated cells. Radiation-induced G2 arrest is prolonged in an acidic environment indicating that the suppression of radiation-induced apoptosis and prolongation of radiation-induced G2 arrest in an acidic environment are related.

The aim was to investigate in detail the influence of intracellular pH (pHi) and intracellular Ca2+ concentration ([Ca2+]i) on apoptosis in HL-60 human promyelocytic leukaemia cells. The pHi was controlled by changing the pH of media as well as by interfering with the pHi regulatory mechanisms with 3-amino-6-chloro-5-(1-homopiperidyl)-N-(diaminomethylene) pyrazincarboxamide (HMA; an inhibitor of Na+/H+ antiport), 4-diiosothiocyanatostilbene-2,2'disulfonic acid, (DIDS; an inhibitor of Na(+)-dependent HCO3-/Cl- exchange) and nigericin (a K+ ionophore). The [Ca2+]i was increased with ionomycin, a Ca2+ ionophore. The apoptosis of HL-60 cells was measured with conventional agarose gel electrophoresis for DNA fragmentation and also with the release of 3H from 3H-thymidine-labelled DNA. Based on the magnitude of DNA fragmentation and 3H release at different pHi, it was shown that apoptosis occurred in HL-60 cells when the pHi was lowered from normal pHi of 7.4 to about 7.2-6.7 with a peak increase at pHi 6.8-6.9. Addition of 4 microM ionomycin to RPMI 1640 medium, which contained 615 microM Ca2+, elevated the apoptosis in the cells. Such an increase in apoptosis by ionomycin in HL-60 cells appeared to result from both an increase in [Ca2+]i and from a decline in pHi. The results indicate that the acidic intratumour environment may greatly affect the response of neoplastic tissues to hyperthermia, radiation and chemotherapeutic drugs which cause apoptosis.

The acidic intracellular environment or low intracellular pH (pHi) increases the thermosensitivity of mammalian cells. The cells in a hypoxic environment produce a greater amount of acidic metabolites than those in an oxygenated environment. However, the hypoxic cells are not more thermosensitive than oxygenated cells since the decrease in pHi is minimized by the mechanisms that regulate the pHi such as Na+/H+ antiport. We hypothesized, therefore, that blocking the regulation of pHi might greatly reduce the pHi and increase the thermosensitivity of hypoxic cells. We tested our hypothesis by heating SCK tumor cells under oxygenated and hypoxic conditions in pH 7.5 or 6.6 medium with or without amiloride, an inhibitor of Na+/H+ antiport. We observed that amiloride increased the thermosensitivity of hypoxic cells markedly. Such a thermosensitization of hypoxic cells by amiloride was more pronounced in an acidic environment, with an enhancement ratio of 1.5, than in a neutral environment, with an enhancement ratio of 1.5. We concluded that lowering the pHi by blocking the regulation of pHi in combination with hyperthermia may be a useful way to eliminate the radioresistant hypoxic cells.

Purpose: The major mechanisms that regulate the intracellular acidity of pHi in mammalian cells are the Na+/H+ exchange and HCO3-/Cl- exchange through the plasma membrane. The purpose of this study was to investigate the feasibility of increasing the thermosensitivity of tumors by increasing intracellular acidity with the use of drugs that inhibit the pHi regulatory mechanisms., Methods and Materials: The pHi of SCK tumor cells in vitro was determined with the fluorescence spectroscopy method. The thermosensitizing effects of the drugs on the cells in neutral (pH 7.2-7.5) and acidic (pH 6.6) media were determined by clonogenic assay. The thermosensitization of SCK tumors in vivo by the drugs was determined with the tumor growth delay and the in vivo-in vitro assay for clonogenic cells., Results: The pHi of SCK tumor cells in pH 7.2-7.5 media was similar to the media pH, while the pHi of the cells in pH 6.6 media was about 7.0. The pHi declined and the thermosensitivity of the tumor cells increased when the Na+/H+ exchange was inhibited with amiloride (3,5 diamino-6-chloro-N-(diaminomethylene) pyrazinecarboxamide) and its analogs, HMA (3-amino-6-chloro-5-(1-homopiperidyl)-N-(diaminomethylene) pyrazinecarboxamide) or EIPA (3-amino-6-chloro-5-(N-ethyl-N-isopropylamino)-N-diaminomethylene) pyrazinecarboxamide), especially in acidic medium. The potencies of HMA and EIPA to decrease the pHi and increase the thermosensitivity in vitro were more than 50 times greater than that of amiloride. DIDS (4,4-diiosothiocyanatostilbene-2,2'-disulfonic acid), an inhibitor of the Na(+)-dependent HCO3-/Cl- exchange, exerted little effect on the pHi and thermosensitivity of SCK cells in vitro, but it enhanced the effects of amiloride and its analogs. Amiloride and HMA also significantly enhanced the thermal effect on tumors in vivo, as judged by the tumor growth delay and also by the in vitro-in vivo assay for clonogenic cells. Combinations of DIDS with amiloride or HMA were more effective than either of them alone in increasing the thermal damage in vivo. As in vitro, HMA was far more potent than amiloride in increasing the thermosensitivity of tumor cells in vivo. However, EIPA was not effective in vivo, probably due to a rapid metabolic breakdown of the drug., Conclusion: The drugs that interfere with the pHi regulatory mechanism significantly thermosensitized the tumor cells in vitro, particularly those in acidic media. The drugs were also effective in increasing the thermosensitivity of tumors. Because the interstitial environment in tumors is acidic relative to that in normal tissues, the thermosensitization by the drugs may be greater in tumors than that in normal tissues.

Purpose: The effects of HMA (3-amino-6-chloro-5-(1-homopiperidyl)-N- (diaminomethylene)pyrazinecarboxamide), an analog of amiloride, on the intracellular pH (pHi) of SCK tumor cells in vitro and on the thermosensitivity of tumors in vivo were investigated., Methods and Materials: The pHi of SCK tumor cells in vitro was measured with the BCEC fluorescence spectroscopy method. The effect of HMA on the thermosensitivity of SCK tumors grown SC in the legs of A/J mice was assessed by the tumor growth delay method and the in vivo-in vitro excision assay method., Results: The pHi of SCK tumor cells in pH 7.5 and 6.6 medium was about 7.50 and 7.15, respectively. The presence of 10-50 microM of HMA lowered the pHi by 0.1-0.2 pH units both in pH 7.5 and 6.6 medium. Heating at 43 degrees C 120 min lowered the pHi by 0.2 and 0.3 pH units in pH 7.5 and 6.6 medium, respectively. When the cells were heated in the presence of 10-50 microM HMA, a marked decline in pHi occurred and and the decline in pHi resulting from the combination of heat and HMA was more pronounced in pH 6.6 medium than in pH 7.5 medium. Heating the SCK tumors grown SC in the legs of A/J mice at 43.5 degrees C for 1 h resulted in a growth delay of 3.7 days. When the host mice were i.v. injected with 0.1 mg/kg of HMA and the tumors were heated heated 20 min later, the tumor growth was delayed by 8.2 days, which was 4.5 days longer than that by heating alone. Heating the SCK tumor at 42.5 degrees C for 1 h caused a tumor growth delay of 0.9 days. An i.v. injection of 1 mg/kg or 10 mg/kg of HMA prior to heating at 42.5 degrees C for 1 h caused a tumor growth delay 2.1 and 3.1 days longer, respectively, than that by heating alone. Such an enhancement of heat-induced tumor growth delay by HMA was due to increased cell killing, as determined with the in vivo-in vitro excision assay of clonogenic cells in the tumors., Conclusion: HMA is a potent thermosensitizer, particularly in an acidic environment. Thermosensitization by HMA may occur preferentially in tumors relative to normal tissues since the intratumor environment is acidic.

It has previously been reported that amiloride, a diuretic drug, sensitizes cells to hyperthermia by inhibiting the Na+/H+ exchange through the plasma membrane and thus decreasing the intracellular pH (pHi), particularly in a low extracellular pH (pHe) environment. In the present study, the efficacy of 5-(N-ethyl-N-isopropyl) amiloride (EIPA), an analog of amiloride, to lower the pHi and sensitize tumor cells to hyperthermia was investigated. It was observed that 10 microM EIPA was as effective as 500 microM amiloride to lower the pHi and to increase the thermal sensitivity of SCK tumor cells in vitro. The fact that lowering the pHi and increasing thermal sensitivity of tumor cells by EIPA are more pronounced in acidic medium suggests that the acidic intratumor environment may be exploited to selectively increase the thermal damage in tumors relative to normal tissues by EIPA or its analogs.

We previously reported that the thermosensitivity of tumor cells can be increased when the intracellular pH is lowered by inhibiting Na+/H+ exchange through the plasma membrane with amiloride (3,5-diamino-6-chloro-N-(diamino methylene)pyrazinecarboxamide) or its analogues and HCO3-/Cl-exchange with 4,4-diiothiocyanato-stilbene-2,2'-disulfonic acid. In this study, we investigated the effects of (3-amino-6-chloro-5- (1-homopiperidyl)-N-(diaminomethylene)pyrazine-carboxamide) (HMA), an analogue of amiloride and a potent inhibitor of Na+/H+ exchange, and R(+)-[(5,6-dichloro-2,3,9,9a-tetrahydro-3-oxo-9a-propyl-1H-fluoren -7- yl)oxy]acetic acid [B-3(+)], a potent inhibitor of HCO3-/Cl- exchange, on the thermosensitivity of SCK tumor cells in vitro. We observed that 10 microM of HMA could effectively increase the cell death by heating at 43 degrees in pH 6.6 medium but not in pH 7.5 medium. The B-3(+) at 50 microM alone had no effect on the thermosensitivity of cells, but it increased the thermosensitizing effect of HMA in acidic medium. Our results strongly suggested that a combination of HMA and B-3(+) may preferentially thermosensitize tumors in vivo since the interstitial environment in tumors is acidic relative to that in normal tissues.

The combined effects of interleukin-1 alpha (IL-1 alpha) and hyperthermia on SCK tumors grown in the legs of A/J mice were investigated. When the host mice were given i.p. injections of 25 micrograms/kg IL-1 alpha, the tumor blood perfusion, as measured with the 86Rb uptake method, significantly declined, reaching minimum blood perfusion in 3-5 h. Although the tumor blood perfusion started to rise thereafter, the recovery was still incomplete 1 day later. Hyperthermia at 42.5 degrees C or 43.5 degrees C for 1 h also caused a marked decline in tumor blood perfusion. When the tumors were heated 5 h after an i.p. injection of IL-1 alpha at 25 micrograms/kg, at which time the tumor blood perfusion was low, the tumor blood perfusion decreased further. The heating of tumors at 42.5 degrees C or 43 degrees C for 1 h significantly reduced the clonogenic cell number in the tumors, delayed the tumor growth, and prolonged the survival time of host animals. The antitumor effect of 25 micrograms/kg IL-1 alpha alone, as judged from the clonogenic cell number, tumor growth delay, and host survival, was smaller than that of hyperthermia. When the host mice were treated with IL-1 alpha and the tumors were heated 5 h later, the decline in the clonogenic tumor cell number, tumor growth delay, and prolongation of host survival were significantly greater than those produced by either treatment alone. It was concluded that the prior reduction of blood perfusion by IL-1 alpha potentiated the antitumor effect of subsequent heatings.

Purpose: Intracellular pH is regulated mainly by Na+/H+ antiport and Cl-/HCO3- exchange through the cell membrane. Amiloride (3,5-diamino-6-chloro-N-(diaminomethylene)pyrazine carboxamide) is a diuretic drug that blocks Na+/H+ antiport and DIDS (4,4-diisothiocyanatostilbene-2,2'-disulfonic acid) is an inhibitor of Cl-/HCO3- exchange. We investigated the potency of these drugs to lower pHi and increase the thermosensitivity of tumors in vivo., Materials and Methods: The cytocidal effect of heat in combination with drug effect in vivo was studied using the in vivo-in vitro clonogenic assay method and the tumor growth delay method with SCK tumors, a mammary adenocarcinoma, on the hind limbs of A/J mice. The effects of amiloride and DIDS on tumor pHi and high energy phosphate levels were investigated using 31P-NMR., Results: We observed that amiloride or DIDS alone increased the effect of hyperthermia at 42.5 degrees C or 43.5 degrees C to suppress tumor growth. The thermosensitization was greater when the two drugs were combined. For example, hyperthermia at 43.5 degrees C alone resulted in a tumor growth delay of about 4 days. When 10 mg/kg amiloride or 25 mg/kg DIDS was injected prior to heating, the growth delay increased to about 6 days. When both drugs were injected prior to heating, a total growth delay of 8 days was obtained. In vivo-in vitro excision assays for cell survival demonstrated that these drugs enhanced the heat-induced tumor cell death. An i.p. injection of 10 mg/kg amiloride plus 25 mg/kg DIDS did not lower the tumor pHi over a 120 min interval. Heating the tumors at 42.5 degrees C for 1 hr significantly lowered the pHi and when the tumor-bearing mice were injected i.p with amiloride and DIDS, and the tumors were heated 1 hr later, the drop in pHi was greater relative to that by heating alone. Heating alone significantly lowered the tumor energy levels as indicated by PCr/Pi and beta-ATP/Pi ratios and an i.p. injection of 25 mg/kg amiloride prior to heating further reduced the energy status in the tumors., Conclusion: Amiloride or its analogs and DIDS may be useful in increasing the therapeutic efficacy of hyperthermia treatments by enhancing the reduction in tumor pHi.

The effects of pentoxifylline (PTX), a drug commonly used for vascular disorders in humans, on the pO2 in SCK tumors of A/J mice and FSa-II tumors of C3Heb/FeJ mice as well as on the radioresponse of SCK tumors were investigated. When the host mice were injected intraperitoneally (ip) with 5 mg/kg PTX, the tumor pO2 increased slowly, peaked 20-50 min postinjection, and returned to its original level in 70-90 min. The magnitude of the increase in tumor pO2 varied markedly depending on the site and tumors. The magnitude of the changes in tumor pO2 after an ip injection of 25 or 50 mg/kg PTX was similar to that caused by 5 mg/kg PTX, but the pO2 tended to remain elevated longer with the higher dose of PTX. When the A/J mice bearing SCK tumors in the legs were injected ip with 50 mg/kg PTX and the tumors were X-irradiated 20 min later, the radiation-induced growth delay of the tumors was greater than that caused by X irradiation alone. The present study demonstrated that PTX is potentially useful for increasing the pO2 and the radioresponse of human tumors.

The effects of amiloride (an inhibitor of Na+/H+ antiport), DIDS (an inhibitor of Na(+)-coupled and Na(+)-independent HCO3-/Cl- exchange) and nigericin (K+/H+ ionophore) alone and in various combinations on the intracellular pH (pHi) and thermosensitivity of SCK tumor cells were studied. Hyperthermia alone at 43 degrees C for 2 h decreased pHi of SCK cells by 0.15-0.20 pH units, as measured fluorometrically using the pH-sensitive dye BCECF. When the cells were treated with 0.5 mM amiloride at 37 degrees C, the pHi declined by 0.10-0.15 pH units at an extracellular pH (pHe) of both 7.2 and 6.6. Amiloride at 0.5 mM enhanced the thermal damage to SCK cells at pHe 6.6 but not at pHe 7.2. DIDS alone at 0.1 mM exerted no effect on pHi or cellular thermosensitivity. DIDS, however, enhanced the effects of amiloride in decreasing pHi and in increasing the thermoresponse of SCK cells, particularly at pHe 6.6. Treatment of the cells with nigericin at 0.1-1.0 micrograms/ml lowered the pHi and enhanced the thermosensitivity of the cells in a dose-dependent manner. Reductions in pHi and increases in thermosensitivity by nigericin at the lower concentration at pHe 6.6 were far greater than at pHe 7.2. When a mixture of 1.0 micrograms/ml nigericin, 0.5 mM amiloride, and 0.1 mM DIDS was present in the medium, the pHi rapidly decreased by about 0.3 and 0.4 pH units at pHe 7.2 and 6.6, respectively. This drug combination was also extremely effective in sensitizing SCK cells to heat, particularly at pHe 6.6. The fact that the thermosensitization by these drugs at pHe 6.6 is more pronounced than at pHe 7.2 and that intratumor environments are known to be acidic strongly suggested that it may then be possible to enhance the thermal damage with such drugs preferentially in tumors relative to normal tissues.

Amiloride, a diuretic drug, is a potent inhibitor of Na+/H+ exchange through the plasma membrane, and has been reported to enhance thermal damage in tumor cells in vitro. We investigated the possible relationship between changes in the thermal response of SCK mouse mammary tumor cells in vitro and changes in intracellular pH (pHi) due to amiloride in the present study. At a concentration of 0.5 mM, amiloride reduced the shoulder (Dq) without causing significant change in the slope (Do) of the survival curve of SCK cells heated once at 43 degrees C. On the other hand, 0.5 mM of amiloride sensitized thermotolerant cells to heat as shown by a reduction in Do. The presence of amiloride during the interval between the first and second heatings slightly reduced or inhibited the development of thermotolerance. The pHi was measured with the BCECF fluorescence method. The presence of 0.5 mM amiloride significantly reduced pHi in both pH 7.2 and 6.6 medium. Heating the SCK cells at 43 degrees C in pH 7.2 or 6.6 medium also reduced the pHi. The combined effect of heat and amiloride in reducing the pHi of SCK cells was additive. These results suggest that the effects of amiloride on the thermal response of SCK cells might be mediated in part by a decrease in pHi. The possibility that the effects of amiloride on the thermal response of tumor cells are mediated by other biochemical changes, such as inhibition of protein synthesis, however, could not be ruled out.

The effectiveness of low extracellular pH in sensitizing cells to heat was studied using SCK mammary carcinoma cells of A/J mice. Solid tumours of 400-600 mg or cells grown in vitro for less than 13 weeks were dispersed to single cells and the in vivo- or in vitro-derived cells were suspended in a medium of pH 7.2 or 5.5-6.6 and then heated at 41-44 degrees C in vitro using a water bath. The dispersion procedure did not alter the heat sensitivity of these cells, but acidic medium of pH below 6.6 caused an increase in heat sensitivity. This effect of low pH decreased with increasing temperature of heating for both cell types. However, the effect was much smaller on in vivo-derived cells than that on in vitro-derived cells for any heating temperature tested. This reduction of the pH effect was also observed for cells derived from larger tumours as well as tumours at an early stage of growth in which the internal milieu was not acidic. This indicates that cellular adaptation to low intratumour pH was not the cause of the reduced pH effect; instead, factors other than low pH must cause the reduced pH effect seen in tumour-derived cells.

The clinical and pathologic features of two brothers with the adult form of Gaucher's disease, both of whom developed cerebral glioblastoma multiforme, are presented. Neither patient had a long-standing neurologic disorder, and morphologic evidence of nervous system glucocerebroside storage was not seen. Despite the known increased incidence of malignancy in Gaucher's disease, a review of the literature and of sixty-two Mayo Clinic cases of adult onset Gaucher's disease revealed only one other instance of the association with glioma. The significance of the relationship is unknown.

Ion-free aluminum hydroxide gel, prepared by the hydrolysis of aluminum tri-sec-butoxide, was observed by IR, X-ray analysis, and pH-stat titration to undergo rapid structural changes leading to the formation of pseudoboehmite and bayerite. The rate of development of order was directly related to the water-aluminum molar ratio. The Al--O bands at 625 and 470 cm-1 were the most sensitive indicators of the development of order in the gel structure. Direct evidence for the stabilizing effect of specifically adsorbed carbonate was obtained when carbon dioxide was introduced during the hydrolysis of aluminum tri-sec-butoxide. The resulting aluminum hydroxycarbonate gel possessed excellent antacid properties, retained its amorphous nature upon aging, and contained no cations other than aluminum. Hydrolysis of aluminum tri-sec-butoxide in the presence of stoichiometric amounts of sodium bicarbonate resulted in the immediate formation of crystalline sodium aluminum hydroxycarbonate (dawsonite).

A retrospective study of 50 surgically confirmed painful hip arthroplasties was performed to identify the roentgenographic findings that are most important in the detection of component loosening or infection. Plain film radiographs and subtraction arthrograms were reviewed in all cases, and specific features revealed by these two radiographic modalities indicative of component failure were identified. Seventeen patients had radioisotope bone scans and 13 of these had additional gallium scans. Plain film roentgenograms correctly identified loosening in 69% of acetabular and 84% of femoral components. Subtraction arthrography had an accuracy of 96% for both acetabular and femoral component loosening. Scintigraphic studies were correct in 77% of acetabular and 89% of femoral components; gallium scans identified 77% of infected hip prostheses.

Herein we describe a previously seldom recognized variety of snapping hip, which is caused by snapping of the iliopsoas tendon over the iliopectineal eminence when the femur is moved from the flexed position at the hip and extended through 45 degrees of flexion. The finding is demonstrated with the patient supine and gently resisting gravity with the iliopsoas muscle, thus placing it under tension. At approximately 45 degrees of flexion, the iliopsoas tendon in two patients was confirmed radiographically to snap abruptly, coincident with an audible sound over the iliopectineal eminence of the pelvis. We believe that this occurs relatively frequently and is generally asymptomatic.