Successful establishment of Wolbachia in Aedes populations to suppress dengue transmission.

Genetic manipulations of insect populations for pest control have been advocated for some time, but there are few cases where manipulated individuals have been released in the field and no cases where they have successfully invaded target populations. Population transformation using the intracellular bacterium Wolbachia is particularly attractive because this maternally-inherited agent provides a powerful mechanism to invade natural populations through cytoplasmic incompatibility. When Wolbachia are introduced into mosquitoes, they interfere with pathogen transmission and influence key life history traits such as lifespan. Here we describe how the wMel Wolbachia infection, introduced into the dengue vector Aedes aegypti from Drosophila melanogaster, successfully invaded two natural A. aegypti populations in Australia, reaching near-fixation in a few months following releases of wMel-infected A. aegypti adults. Models with plausible parameter values indicate that Wolbachia-infected mosquitoes suffered relatively small fitness costs, leading to an unstable equilibrium frequency <30% that must be exceeded for invasion. These findings demonstrate that Wolbachia-based strategies can be deployed as a practical approach to dengue suppression with potential for area-wide implementation.

The Prevalence of Clinically Significant Prostate Cancer According to Commonly Used Histological Thresholds in Men Undergoing Template Prostate Mapping Biopsies.

PURPOSE: Transrectal prostate biopsies are inaccurate and, thus, the prevalence of clinically significant prostate cancer in men undergoing biopsy is unknown. We determined the ability of different histological thresholds to denote clinically significant cancer in men undergoing a more accurate biopsy, that of transperineal template prostate mapping. MATERIALS AND METHODS: In this multicenter, cross-sectional cohort of men who underwent template prostate mapping biopsies between May 2006 and January 2012, 4 different thresholds of significance combining tumor grade and burden were used to measure the consequent variation with respect to the prevalence of clinically significant disease. RESULTS: Of 1,203 men 17% (199) had no previous biopsy, 38% (455) had a prior negative transrectal ultrasound biopsy, 24% (289) were on active surveillance and 21% (260) were seeking risk stratification. Mean patient age was 63.5 years (SD 7.6) and median prostate specific antigen was 7.4 ng/ml (IQR 5.3-10.5). Overall 35% of the patients (424) had no cancer detected. The prevalence of clinically significant cancer varied between 14% and 83% according to the histological threshold used, in particular between 30% and 51% among men who had no previous biopsy, between 14% and 27% among men who had a prior negative biopsy, between 36% and 74% among men on active surveillance, and between 47% and 83% among men seeking risk stratification. CONCLUSIONS: According to template prostate mapping biopsy between 1 in 2 and 1 in 3 men have prostate cancer that is histologically defined as clinically significant. This suggests that the commonly used thresholds may be set too low.

GDF15 is a potential predictive biomarker for TPF induction chemotherapy and promotes tumorigenesis and progression in oral squamous cell carcinoma.

BACKGROUND: Randomized trials have not shown major survival benefits when induction chemotherapy plus standard therapy is compared with standard therapy alone in patients with oral squamous cell carcinoma (OSCC). Induction chemotherapy is likely to be effective for biologically distinct subgroups and biomarker development may lead to identification of patients whose tumors are likely to respond to a particular treatment. PATIENTS AND METHODS: We evaluated immunohistochemical staining for GDF15 in pretreatment biopsy specimens of 230 of 256 OSCC patients who were treated in a prospective, randomized, phase III trial on induction chemotherapy including docetaxel, cisplatin and 5-fluorouracil (TPF). Relationship between GDF15 intervention and cell proliferation, migration, invasion, colony formation and tumorigenicity was analyzed using in vitro and in vivo OSCC models. RESULTS: Low GDF15 expression predicted a better survival in OSCC patients, especially overall survival [P = 0.049, hazard ratio (HR) = 0.597] and distant metastasis-free survival (DMFS; P = 0.031, HR = 0.562). cN+ patients with low GDF15 expression benefitted from induction TPF in overall survival (P = 0.039, HR = 0.247) and DMFS (P = 0.039, HR = 0.247), cN- patients with high GDF15 expression benefitted from induction TPF in overall survival (P = 0.019, HR = 0.231), disease-free survival (P = 0.011, HR = 0.281), locoregional recurrence-free survival (P = 0.035, HR = 0.347) and DMFS (P = 0.009, HR = 0.197). Decreased GDF15 expression in OSCC lines significantly inhibited cell proliferation, migration, invasion, colony formation and tumorigenesis through increased phosphorylation of AKT and ERK1/2 (P < 0.05). Likewise, overexpression of GDF15 significantly promoted cell proliferation, migration, invasion and colony formation through decreased phosphorylation of AKT and ERK1/2 (P < 0.05). CONCLUSIONS: GDF15 expression can be used as a prognostic biomarker for OSCC, and as a predictive biomarker for benefitting from TPF induction chemotherapy. GDF15 promotes tumorigenesis and progression through phosphorylation of AKT and ERK1/2 in OSCC. The clinical trial in this study was registered with www.ClinicalTrials.gov (NCT01542931).

Organizational Leaders Perceptions of Barriers to Accessing Behavioral Health Services in a Low-Resource Community.

Little is known about how to effectively implement behavioral health programs in low-resource communities. Leaders from 20 community-serving behavioral health organizations in Flint, MI, were asked about their organizations and the barriers that they, and the populations they serve, face in providing and accessing behavioral health services. Barriers are reported using a mixed-methods analysis, reporting the number and percentage of organizations that experienced the barrier along with example quotations from the organization leaders. The most frequently reported barrier to providing services was finding adequate funding (50%) while the most frequently reported barrier for accessing services was finding adequate and reliable transportation (30%). Comparisons of these findings with barriers reported by providers in different settings and those seeking services are discussed. These comparisons may provide an important next step in identifying areas where providers perceptions and the needs of the population are misaligned and for systemic improvements more broadly.

Evaluation of the upper urinary tract using transureteric ultrasound--a review of the technique and typical imaging appearances.

Ureteric strictures and pelviureteric junction obstruction often present a diagnostic conundrum to radiologists, particularly after the first-line investigations have failed to provide a definitive answer. Transureteric ultrasonography (TUU) is a relatively novel technique performed by the radiologist, which uses a miniature endoluminal ultrasound probe to interrogate the ureteric anatomy and peri-ureteric soft tissues. In this review, we discuss how TUU is performed, and the normal imaging appearances of the ureter and surrounding anatomical structures. We also focus on the various pathological processes that can be accurately evaluated or diagnosed using TUU including lymphadenopathy, calculi, ureteric neoplasms, ureteritis, crossing vessels and aneurysms. As TUU is not well established in UK practice as yet, we suggest possible indications for its use in the diagnostic work-up of urological patients and future applications.

Tumor-promoting phorbol ester down-regulates the androgen induction of prostate-specific antigen in a human prostatic adenocarcinoma cell line.

Prostate-specific antigen (PSA) is the most sensitive marker available for monitoring the progression of prostate cancer and response to therapy. In a previous study, we demonstrated tissue-specific expression of PSA glycoprotein and mRNA and its regulation through the androgen receptor. In this study, we examine the effects of protein kinase A (PKA) and protein kinase C (PKC) on the androgen regulation of PSA in a human adenocarcinoma cell line, LNCaP. Northern blot analysis demonstrated that forskolin, an activator of PKA, had no effect on the androgen regulation of PSA. However, the phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA), a direct activator of PKC, showed a time- and dose-dependent repression of the androgen regulation of PSA glycoprotein and mRNA. The biologically inactive phorbol ester, 4 alpha-phorbol-12,13-didecanoate, had no effect. Staurosporine, a PKC inhibitor, blocked the TPA-mediated repression of the androgenic stimulation of PSA glycoprotein. In addition, the calcium ionophore, A23187, was able to simulate the actions of TPA, presumably through activation of PKC via calcium mobilization. In summary, the androgenic regulation of PSA protein and mRNA is repressed by tumor-promoting phorbol esters through the PKC pathway. This indicates that the effects of TPA may be secondary to repressed gene transcription or altered mRNA stability. In addition, this study emphasizes that the androgenic regulation of PSA is complex and may involve other extracellular transduction signals.

Hormonal regulation of prostate-specific antigen messenger RNA in human prostatic adenocarcinoma cell line LNCaP.

Prostate-specific antigen (PSA) is a member of the kallikrein gene family and is expressed exclusively in human prostatic epithelial cells. PSA protein has been an important biological marker for prostate cancers. Until now, very little was known about the regulation of PSA expression in prostatic cells. In this study, we have developed a specific oligonucleotide probe which recognizes PSA but not the human glandular kallikrein. This is crucial because both PSA and human glandular kallikrein are expressed in the prostate at relatively high levels and have high nucleotide sequence homology (greater than 82%). Utilizing a S-labeled PSA-specific probe, PSA mRNA was localized within the glandular epithelium of the prostate. Northern blot analysis detected a single 1.6-kilobase transcript in LNCaP cells, a cell line derived from a human prostate adenocarcinoma metastasis. Therefore, LNCaP cells were used to study the androgenic effects on PSA mRNA expression. A time course study demonstrated that PSA mRNA was induced by mibolerone (a nonmetabolizable synthetic androgen) and reached maximal levels after 9 h. The induction of PSA mRNA required as little as 0.3 nM mibolerone. In addition to mibolerone, PSA mRNA could be induced by the natural androgen, dihydrotestosterone, but not by the synthetic glucocorticoid, dexamethasone, or the synthetic estrogen, diethylstilbestrol. Moreover, in the presence of dihydrotestosterone, PSA mRNA was depressed by hydroxyflutamide (an antiandrogen). These results suggest strongly that the androgenic effects on PSA mRNA in LNCaP cells may be via the function of the androgen receptor.

Does needle size matter? Patient experience of luteinising hormone-releasing hormone analogue injection.

To determine whether needle size influences a patient's perception of pain, 50 patients requiring hormonal manipulation for prostate cancer were blindfolded and randomised to receive two goserelin ('Zoladex') or two leuprorelin ('Prostap') injections, using 16- or 23-gauge needles, respectively. Median visual analogue scale pain scores for the first injections of goserelin and leuprorelin were below the level of clinical significance and were not statistically different. Mean administration time for goserelin was significantly shorter than for leuprorelin. In conclusion, there was no statistically significant difference in pain experienced on injection of goserelin and leuprorelin when patients were unaware of needle size.

Glucocorticoids decrease interleukin-6 levels and induce mineralization of cultured osteogenic cells from children with fibrous dysplasia.

Fibrous dysplasia (FD) is a progressive bone disease in which abnormal fibroblast proliferation results in the replacement of normal cancellous bone with an immature fibrous tissue that is poorly mineralized. The disease manifests itself in the monostotic form in which only one bone is involved and the polyostotic form in which multiple bones at different sites are affected. The McCune-Albright syndrome is a variation of the polyostotic form in which patients demonstrate a greater extent of bone involvement and a variety of endocrinopathies. Somatic activating mutations in the GNAS gene have been demonstrated in the fibrotic lesions of patients affected with either monostotic or polyostotic FD. The increased cAMP levels caused by the G-protein mutations lead to increased interleukin-6 (IL-6) levels in the affected tissues, resulting in abnormal osteoblast differentiation and increased osteoclastic activity. Utilizing cell culture techniques that have been developed for mammalian bone marrow stromal cells, we have successfully cultured osteogenic stem cells from the affected stroma of 11 FD patients. Cells cultured from patients with polyostotic FD showed a high frequency of the Gsalpha mutation, whereas cells from monostotic FD patients showed a low frequency of the mutation. Both the normal and FD cells displayed the osteogenic phenotype when exposed to medium containing glucocorticoids. Glucocorticoids also caused a dramatic inhibition of IL-6 mRNA and protein levels in osteogenic cells cultured from the FD patients. These findings suggest that chemical alteration of cellular function may lead to new treatment options for patients with FD.

Importance of early phase insulin secretion to intravenous glucose tolerance in subjects with type 2 diabetes mellitus.

Insulin secretion is impaired in type 2 diabetes with the early response being essentially absent. The loss of this early insulin secretion is hypothesized to be important in the deterioration of glucose tolerance. To determine whether enhancement of the early-phase insulin response can enhance glucose tolerance, we administered 1) 120 mg nateglinide, an insulinotropic agent that enhances early insulin secretion; 2) 10 mg glyburide, which enhances the later phases of insulin secretion; or 3) placebo in random order to 21 subjects with type 2 diabetes (14 males and 7 females; aged 59.2 +/- 2.1 yr, x +/- SEM; body mass index 29.7 +/- 1.0 kg/m(2); fasting plasma glucose 8.1 +/- 0.1 mM). beta-Cell function was quantified as the incremental area under the curve for different time periods for the 5 h following iv glucose administration and glucose tolerance as the glucose disappearance constant (Kg) from 10 to 60 min. Insulin release commenced immediately after nateglinide administration, even before glucose injection, but this was not observed with glyburide. Both nateglinide and glyburide enhanced glucose-induced insulin release, compared with placebo (area under the curve -15-300 min: nateglinide 23,595 +/- 11,212 pM/min, glyburide 54,556 +/- 15,253 pM/min, placebo 10,242 +/- 2,414 pM/min). The profiles of insulin release demonstrated significant enhancement of release between -15 and 30 min for nateglinide, compared with glyburide and between 60 and 300 min for glyburide over nateglinide. Kg increased by 15% with nateglinide (0.87 +/- 0.04%/min), but it did not increase significantly with glyburide (0.79 +/- 0.04%/min), compared with placebo (0.76 +/- 0.04%/min). The enhancement of insulin release by glyburide resulted in a lower minimal glucose concentration with glyburide (3.8 +/- 0.2 mM), compared with nateglinide (5.0 +/- 0.2 mM) and placebo (5.9 +/- 0.2 mM). Thus, enhancement of the early phase of insulin secretion improves iv glucose tolerance, whereas delaying it by 30 min results in a slower rate of glucose disappearance for the first 2 h after iv glucose administration. Further, the differences in the kinetics of nateglinide and glyburide action results in continued insulin release with glyburide despite the fact that glucose levels have returned to basal, thus resulting in a further reduction in glucose levels and a lower nadir.

Age as a function of ATPase activity in cultured corneal endothelial cells.

As cells grown in tissue culture age, they typically become altered when compared to their in vivo counterparts. Bovine corneal endothelial cells were grown in culture for periods up to 60 days (primary = 10 days; secondary = 50 days). The plasma membrane enzymes: Na+K+ ATPase and Mg2+ ATPase were assayed for specific activity at selected intervals throughout the growth periods. In primary cultures, it was found that Na+K+ ATPase was quite low at 5 days (0.05 units), but increased fourfold at 10 days (0.22 units). By contrast, Mg2+ ATPase changed little over the same period. Tissue cultures grown secondarily had Na+K+ ATPase activity fall 0.3 units (0.52 to 0.22) for 35 days after a single trypsinization. After five trypsinizations over a 50-day period, the activity fell 0.23 units (0.33 to 0.10). The alterations in Mg2+ ATPase activity were more complex in secondary cultures. In the instance in which a single trypsinization was used, the activity fell 0.15 units at day 20, rose 0.12 units (above the starting value) at day 25, then returned to the initial level by day 35. When multiple trypsinizations were used, the activity fell 0.06 units by day 30, then remained stable for the duration. The data may be indicative of mechanisms such as receptor alteration, feedback inhibition and genetic instability when these cells are grown in culture.

Relationship of telomeres and p53 in aging bovine corneal endothelial cell cultures.

PURPOSE: To demonstrate a relationship between telomere lengths and levels of p53 in cultured bovine corneal endothelial cells (CECs) during aging. METHODS: Bovine CECs were grown and aged as long-term cultures. Telomere lengths were determined directly on gels with 32P probes after treatment of isolated DNA with RsaI and HinfI. Protein p53 was determined using an enzyme-linked immunosorbent sandwich assay. Cellular aging and the development of replicative senescence were monitored by the appearance of senescent morphology and the beta-galactosidase assay. RESULTS: Bovine CEC telomeres lost 4 kb (from 12.8 to 8.8 kb) over 1 year (89 population doublings [PDs]). The p53 levels in bovine CECs were initially small (approximately 60 pg/million cells), but rose 3.5-fold by culture age of 260 days (64 PDs). On initiation, cultured bovine CECs did not stain for the senescent marker beta-galactosidase. However, these cells stained at 89 PDs and senescent morphology was observed in the cultures at 64 PDs. CONCLUSIONS: The data indicate an inverse relationship between telomere lengths (decreasing) and levels of p53 (increasing) in bovine CECs during aging. These properties may influence the ability of these cells to divide as they enter into replicative senescence.

Stage B prostate adenocarcinoma. Flow cytometric nuclear DNA ploidy analysis.

Over a 16-year period (1966 to 1981), 349 patients underwent radical retropubic prostatectomy for pathologic stage B adenocarcinoma of the prostate. Nuclear DNA content was measured by flow cytometry on available archival material of 283 patients. Two hundred sixty-one patients (92%) had high-quality histograms. The ploidy distribution was as follows: DNA diploid, 177 (68%); DNA tetraploid, 74 (28%); and DNA aneuploid, 10 (4%). The average follow-up was 9.4 years. At the time of follow-up, 53 patients (20%) within the study group had developed tumor progression: 22 local, 23 systemic, and 8 both. The ploidy distribution of the population that developed tumor progression was 27 DNA diploid (51%), 16 DNA tetraploid (30%), and 10 DNA aneuploid (19%). This ploidy distribution is significantly different from that found for the nonprogression group with stage B disease. Overall, 31% of patients with DNA nondiploid tumors had tumors that progressed compared with 15% of patients with DNA diploid tumors. All (100%) DNA aneuploid tumors progressed. The DNA ploidy distribution of all pathologic stage B prostate cancers differs significantly from that found in more advanced stages (C and D1) previously reported for the same time interval. However, the ploidy distribution of stage B tumors that progressed closely resembles that of the stage C and D1 tumors. These results further support the working hypothesis that nuclear DNA content has marked prognostic significance for patients with adenocarcinoma of the prostate. It seems to us that analysis of ploidy by flow or static cytometry will become an essential tool for treating patients with localized prostate cancer.

Outcome analysis and patient satisfaction following octant transrectal ultrasound-guided prostate biopsy: a prospective study comparing consultant urologist, specialist registrar and nurse practitioner in urology.

PURPOSE: To determine whether transrectal ultrasound-guided biopsy of the prostate is equally reliable and acceptable if performed by urology nurse practitioner or urologist. SCOPE: Octant biopsies were taken by each operator (consultant urologist n=2, urology specialist registrar n=1 and urology nurse practitioner n=2) from 50 consecutive unselected patients and demographics and cancer detection rate were compared between the groups. A postal survey was performed following nurse practitioner biopsy to assess patient satisfaction and acceptance of nurse practitioner biopsy. CONCLUSION: Transrectal ultrasound-biopsy of prostate whether performed by nurse practitioner or urologist is equally reliable if adequate training is provided. Patients are happy to undergo prostate biopsy and receive information about the diagnosis from an appropriately trained prostate cancer nurse specialist.

Pain, coping, and adjustment in patients with burns: preliminary findings from a prospective study.

We prospectively examined the associations between procedural pain during hospitalization and coping and adjustment 1 month postdischarge in 43 patients treated at a major regional burn center for burns extensive enough to require at least 5 days of daily wound debridement procedures. Both patients and nurses provided ratings of patient pain, which were summarized and aggregated across a 5-day period. Results indicated that those subjects with higher pain scores also reported poorer adjustment as measured by scores on the Brief Symptoms Inventory and the Sickness Impact Profile. Moreover, these associations remained significant after partialling out the effects of preburn adjustment. Hierarchical regression analyses revealed evidence that seeking social support had a moderating effect on the association between pain and scores on a measure of posttraumatic stress disorder.

Effects of mannitol on cultured corneal endothelial cell Na,K-ATPase activity.

Mannitol was introduced into the media of bovine corneal endothelial cells grown in culture. This was accomplished in order to compare its influence on Na,K-ATPase activity with that of high levels of glucose that inhibit the enzyme. The study was conducted with the intent of showing possible adverse osmolar effects on enzyme activity. Mannitol was found to inhibit NA,K-ATPase when compared with mannitol-free medium (11 U vs. 202 U). However, this inhibition was significantly greater than that produced by high levels of glucose. When mannitol was introduced directly into the assay for the plasma membrane-extracted enzyme, the inhibition was as severe (14 U) as for that placed in the culture medium. By way of comparison, glucose introduced into the enzyme assay caused no significant inhibition (189 U). The mannitol concentration used was 20 mM and in the media there was an osmotic pressure of 347 mOsmol/kg. The high glucose concentration was 25 mM and the osmotic pressure in the media was 341 mOsmol/kg. These osmotic pressures were compared with that of the control medium (with 5 mM glucose), which generated a pressure of 308 mOsmol/kg. None of these values were judged sufficiently high to rupture cell plasma membranes or alter cell morphology as seen by vital staining and phase contrast microscopy. In addition, it was found that mannitol had no effect on cellular DNA, whereas a previous study showed that high glucose caused an increased unwinding of duplex DNA. This study suggests that mannitol inhibits endothelial cell Na,K-ATPase by a different mechanism than that of glucose. It further points out that osmotic effects may not be involved with either mechanism.

Alteration of ATPase activity and duplex DNA in corneal cells grown in high glucose media.

An investigation was made on the possible effects of high levels (450 mg/dl) of glucose on the activities of Na,K-ATPase [E.C. (Enzyme Commission) 3.6.1.37] and Mg-ATPase (E.C. 3.6.1.4) in plasma membrane preparations of bovine corneal endothelial cells grown in tissue culture. The activities of these enzymes were compared with the activities of the same enzymes from cells grown in low or "fasting" levels (90 mg/dl) of glucose. All activities were assayed from cells that were secondary cultures (15-25 days after trypsinization). The results indicated a 76% decrease in activity for Na,K-ATPase (0.04 units in high glucose vs. 0.17 units in low glucose). The activity for Mg-ATPase also decreased by 33% (0.24 units in high glucose vs. 0.36 units in low glucose). A t-test for significance indicated that the loss of activity in both enzymes was highly significant (p < 0.001) in the high glucose media. Assays for ATPase activity of plasma membranes were also made directly in high glucose after removal of the membranes from cells grown in low-glucose media. However, those membrane ATPases showed no significant decrease in activity. Tests for DNA damage of cells grown in the presence of high levels of glucose indicated a 15.5% change (decrease) in the amount of double-stranded DNA remaining after alkali treatment. This change was highly significant also (p < 0.001). These data suggest that the diabetic state may negatively affect membrane-bound ATPases of the corneal endothelium and further point to the possibility of an altered synthetic rate of ATPase polypeptides as a result of DNA damage.

The acidification response of normal subjects to ammonium chloride using a 3-day loading test.

The acidification response to NH4 Cl loading (0.1 g/kg bw/day) was tested in 16 normal healthy subjects in the basal fasting state on Day 4, the subjects having taken the salt daily for the 3 previous days. The response was measured in terms of blood pH and in urine, creatinine, phosphate, pH, titratable acidity, ammonium, net acidity and creatinine clearance. To minimise inter-subject variation the urine values were adjusted to a standard body surface area of 1.73 m2. A normal range for the blood pH of the mean value +/- 2 SD, encompassed the observed range of values. However, to fit the observed range of acid-base values in urine into the 2 SD range required a logarithmic transformation of the data. Statistical analysis confirmed a significant correlation between blood [H+] net acid secretion, urine titratable acidity and ammonium. Urine net acid secretion was positively correlated with urinary phosphate, titratable acidity and ammonium.