Air pollution accountability research: Moving from a chain to a web.

Air pollution accountability studies examine the relationship(s) between an intervention, regulation, or event and the resulting downstream impacts, if any, on emissions, exposure, and/or health. The sequence of events has been schematically described as an accountability chain. Here, we update the existing framework to capture real-life complexities and to highlight important factors that fall outside the linear chain. This new "accountability web" is intended to convey the intricacies associated with conducting an accountability study to various audiences, including researchers, policy makers, and stakeholders. We also identify data considerations for planning and completing a robust accountability study, including those relevant to novel and innovative air pollution and exposure data. Finally, we present a series of recommendations for the accountability research community that can serve as a guide for the next generation of accountability studies.

"Mothers get really exhausted!" The lived experience of pregnancy in extreme heat: Qualitative findings from Kilifi, Kenya.

Heat exposure in pregnancy is associated with a range of adverse health and wellbeing outcomes, yet research on the lived experience of pregnancy in high temperatures is lacking. We conducted qualitative research in 2021 in two communities in rural Kilifi County, Kenya, a tropical savannah area currently experiencing severe drought. Pregnant and postpartum women, their male spouses and mothers-in-law, community health volunteers, and local health and environment stakeholders were interviewed or participated in focus group discussions. Pregnant women described symptoms that are classically regarded as heat exhaustion, including dizziness, fatigue, dehydration, insomnia, and irritability. They interpreted heat-related tachycardia as signalling hypertension and reported observing more miscarriages and preterm births in the heat. Pregnancy is conceptualised locally as a 'normal' state of being, and women continue to perform physically demanding household chores in the heat, even when pregnant. Women reported little support from family members to reduce their workload at this time, reflecting their relative lack of autonomy within the household, but also potentially the 'normalisation' of heat in these communities. Climate change risk reduction strategies for pregnant women in low-resource settings need to be cognisant of local household gender dynamics that constrain women's capacity to avoid heat exposures.

Complications and satisfaction in transwomen receiving breast augmentation: short- and long-term outcomes.

BACKGROUND: To achieve long-term improvement in health care of transgender women, it is necessary to analyze all aspects of gender-confirming surgery, especially the relation of risks and benefits occurring in these procedures. While there are many studies presenting data on the urologic part of the surgery, there are just few data about complications and satisfaction with breast augmentation. METHODS: This is a retrospective study using parts of the BREAST-Q Augmentation Questionnaire and additional questions for symptoms of capsular contracture and re-operations and analyzing archived patient records of all transwomen which were operated at University Hospital Essen from 2007 to 2020. RESULTS: 99 of these 159 patients (62%) completed the questionnaire after a median time of 4 years after surgery. Breast augmentation led to re-operations due to complications in 5%. The rate of capsular contracture (Baker Grad III-IV) in this population was 3%. Most patients (75%) rated high scores of satisfaction with outcome (more than 70 points) and denied to have restrictions due to their implants in their everyday life. All patients reported an improvement in their quality of life owing to breast augmentation. CONCLUSION: Breast augmentation by inserting silicon implants is a safe surgical procedure which takes an important part in reducing gender dysphoria.

Biopsy-derived oral keratinocytes - A model to potentially test for oral mucosa radiation sensitivity.

Purpose: To establish stable in vitro growth of keratinocytes from very small biopsy specimens and successfully apply new test systems to determine their radiosensitivity. Materials and Methods: Oral mucosa biopsies (diameter: 1.7 mm) from 15 subjects were immobilized with custom-made cups onto culture plates. Outgrowing cells were tested for cytokeratin 5/14 and Ki67, expanded, radiated at different doses, and seeded onto circumscribed areas before being allowed to spread centrifugally. In this newly developed spreading assay, cell-covered areas were measured by image analysis. For statistical analysis, a linear mixed regression model was used; additionally, results were correlated to the radiation dose applied. Colony forming efficiency (CFE) was used to validate the results. DNA damage repair was analysed by gammaH2AX and 53BP1 foci quantification using immunofluorescence microscopy 24 h and 96 h after irradiation. Results: Stable keratinocyte growth continued for up to 7 weeks in 14 biopsies. Cells spread reliably from an initial 16.6 mm2 up to a median of 119.2 mm2 (range: 54.4-290). Radiated cells spread to only 100.7 mm2 (2 Gy; range: 55.3-266.7); 73.2 mm2 (4 Gy; 15-240.4); 47 mm2 (6 Gy; 2-111.9), and 22.7 mm2 (8 Gy; 0-80). Similarly, CFE decreased from 0.223 (0 Gy) to 0.0028 (8 Gy). Using an individual donor as a random factor, cell spread correlated with CFE, where radiation dose was the main driver (decrease by 0.50, adjusted for area). Upon irradiation with 6 Gy, radiation-induced DNA damage was increased after 24 h in all samples, and even after 96 h in 5 out of 7 samples, as detected by a higher number of gammaH2AX/53BP1 foci in irradiated cells (mean 3.7 for 24 h; mean 0.6 for 96 h). Conclusion: In vitro propagation of keratinocytes derived from a small biopsy is feasible. Radiation impairs cellular migration and proliferation, and the newly described spreading assay allows ranking for cellular radioresistance. The keratinocyte model also supports classical functional assays such as clonogenic survival and DNA double strand repair. The clinical relevance awaits upcoming investigations.

Efficacy, safety and quality-of-life data from patients with pre-treated metastatic colorectal cancer receiving trifluridine/tipiracil: results of the TALLISUR trial.

INTRODUCTION: Trifluridine/tipiracil (FTD/TPI) improved both overall and progression-free survival (OS, PFS) of patients with pre-treated metastatic colorectal cancer (mCRC) in the pivotal phase III RECOURSE trial. However, health-related quality of life (HRQoL) was not assessed directly. To this end and to generate post-authorisation data, the TALLISUR trial was conducted. METHODS: In this prospective, multi-centre, Germany-wide, phase IV study, patients with pre-treated mCRC were given the choice to receive either FTD/TPI or best supportive care (BSC). A validated questionnaire, EORTC QLQ-C30, was employed to assess HRQoL. Secondary endpoints included OS, PFS and safety. RESULTS: Of 194 eligible patients, 185 decided to receive FTD/TPI and 9 to receive BSC. The low number of patients in the BSC-arm did not allow statistically meaningful analyses. On the other hand, treatment with FTD/TPI was associated with maintained HRQoL. Median OS was 6.9 months [95% confidence interval (CI) 6.1-8.2 months] and median PFS was 2.5 months (95% CI 2.1-2.9 months). The most frequent treatment-emergent adverse events were neutropenia (27.6%) and anaemia (22.7%). Febrile neutropenia occurred in 1.1%. CONCLUSIONS: Treatment of patients suffering from pre-treated mCRC with FTD/TPI was associated not only with prolonged survival and delayed progression but also with maintained HRQoL.

Comparative analysis of Bricker versus Wallace ureteroenteric anastomosis and identification of predictors for postoperative ureteroenteric stricture.

PURPOSE: Ureteroenteric anastomosis after cystectomy is usually performed using the Bricker or Wallace technique. Deterioration of renal function is the most common long-term complication of urinary diversion (UD). To improve surgical care and optimize long-term renal function, we compared the Bricker and Wallace anastomotic techniques and identified risk factors for ureteroenteric strictures (UES) in patients after cystectomy. MATERIAL AND METHODS: Retrospective, monocentric analysis of 135 patients who underwent cystectomy with urinary diversion at the University Hospital Essen between January 2015 and June 2019. Pre- and postoperative renal function, relevant comorbidities, prior chemo- or radiotherapy, pathological findings, urinary diversion, postoperative complications, and ureteroenteric strictures (UES) were analyzed. RESULTS: Of all 135 patients, 69 (51.1%) underwent Bricker anastomosis and 66 (48.9%) Wallace anastomosis. Bricker and Wallace groups included 134 and 132 renal units, respectively. At a median follow-up of 14 (6-58) months, 21 (15.5%) patients and 30 (11.27%) renal units developed UES. We observed 22 (16.6%) affected renal units in Wallace versus 8 (5.9%) in Bricker group (p < 0.001). A bilateral stricture was most common in Wallace group (69.2%) (p < 0.001). Previous chemotherapy and 90-day Clavien-Dindo grade ≥ III complications were independently associated with stricture formation, respectively (OR 9.74, 95% CI 2-46.2, p = 0.004; OR 4.01, 95% CI 1.36-11.82, p = 0.013). CONCLUSION: The results of this study show no significant difference in ureteroenteric anastomotic techniques with respect to UES development regarding individual patients but suggest a higher risk of bilateral UES formation in patients undergoing Wallace anastomosis. This is reflected in the increased UES rate under consideration of the individual renal units.

Transrectal Ultrasound Guided Hydrodistension - A New Surgical Way in Transgender Surgery.

BACKGROUND: Creating the neovaginal canal in transwomen is one of the most delicate steps of Genital Gender Affirming Surgery (GGAS). Injury to the rectum is a rare but serious complication that can lead to further surgery and even creation of a colostomy. AIM: Implementation of a novel hydrospacing technique (HST) based on transrectal ultrasound (TRUS)-guided hydrodistension. METHODS: Between June 2018 and June 2020 54 transwomen received GGAS with HST. Immediately before GGAS transperineal hydrodistension was performed using a TSK-Supra-Needle (20 Gauge, 120 mm length), that was placed under direct TRUS-guided visual control between Denonvilliers' fascia and the anterior rectal wall. 40 - 60 ml normal saline were administered perineally to separate Denonvilliers' fascia from the anterior rectal wall to create a dissection of at least 20 mm. For better intraoperative visualization the hydrodissected space was also dyed using 2ml of methylenblue while retracting the needle. A retrospectively analysed, clinically and demographically comparable series of 84 transwomen who underwent GGAS between June 2016 and June 2018 served as control group. All 138 surgeries were performed by the same experienced surgeon. OUTCOMES: The effect of the novel hydrospacing technique on neovaginal dimensions and operating time. RESULTS: Patients in both groups did not differ in baseline patient characteristics such as age and body mass index (HST 35 vs 38 years in control group, P = .44 and body mass index 26 vs 25 kg/m2, P = .73). Vaginal depth and width were significantly larger in the HST subgroup as compared to controls (14.4 cm vs 13.5 cm, P = .01 and 4.2 cm vs 3.8 cm, P < .001). No statistically significant difference occurred in intraoperative rectal injury (n = 0 in HST group, n = 2 in control group, P = .26). Median total OR-time was comparable for GGAS including HST before vaginoplasty to standard technique (211 minutes for HST vs 218 minutes; P = 0.19). CLINICAL IMPLICATIONS: The proposed additional surgical step during GGAS is minimally invasive and safe, simplifies GGAS and potentially helps to avoid complications such as rectal injury. STRENGTH & LIMITATIONS: Single-surgeon series, limited follow-up time and no prospective randomization. CONCLUSION: HST is a safe and feasible procedure, which facilitates a safe preparation of the neovaginal canal during male to female GGAS. Panic A, Rahmani N, Kaspar C, et al. Transrectal Ultrasound Guided Hydrodistension - A New Surgical Way in Transgender Surgery. J Sex Med 2021;18:1135-1141.

A systematic review of the effects of temperature and precipitation on pollen concentrations and season timing, and implications for human health.

Climate and weather directly impact plant phenology, affecting airborne pollen. The objective of this systematic review is to examine the impacts of meteorological variables on airborne pollen concentrations and pollen season timing. Using PRISMA methodology, we reviewed literature that assessed whether there was a relationship between local temperature and precipitation and measured airborne pollen. The search strategy included terms related to pollen, trends or measurements, and season timing. For inclusion, studies must have conducted a correlation analysis of at least 5 years of airborne pollen data to local meteorological data and report quantitative results. Data from peer-reviewed articles were extracted on the correlations between seven pollen indicators (main pollen season start date, end date, peak date, and length, annual pollen integral, average daily pollen concentration, and peak pollen concentration), and two meteorological variables (temperature and precipitation). Ninety-three articles were included in the analysis out of 9,679 articles screened. Overall, warmer temperatures correlated with earlier and longer pollen seasons and higher pollen concentrations. Precipitation had varying effects on pollen concentration and pollen season timing indicators. Increased precipitation may have a short-term effect causing low pollen concentrations potentially due to "wash out" effect. Long-term effects of precipitation varied for trees and weeds and had a positive correlation with grass pollen levels. With increases in temperature due to climate change, pollen seasons for some taxa in some regions may start earlier, last longer, and be more intense, which may be associated with adverse health impacts, as pollen exposure has well-known health effects in sensitized individuals.

M0CRPC overview of management options.

Though prostate cancer usually responds to androgen deprivation therapy (ADT) in the beginning, the majority of prostate cancers will develop castration resistance over time. The androgen receptor (AR) pathway is often found to be activated in castration resistant prostate cancer (CRPC). Thus, AR signalling remains a therapeutic target upon the development of CRPC. The term M0CRPC is used when ADT leads to castration resistance and there are no metastases detectable by means of conventional imaging. Until recently, there was no therapeutic standard for this group of patients. With the PROSPER-, SPARTAN- and ARAMIS-studies three large placebo-controlled phase III trials have been published lately that showed a significant benefit in metastasis-free survival in men with M0CRPC and short PSA doubling time (PSADT). The efficacy data are very similar in these studies, meaning that the drugs' safety profiles, final analyses of overall survival and their availability will be more important to help clinicians decide which of these three drugs they use for their patients with M0CRPC.

Sex disparities determined by biometric and hematological measurements in Astyanax bimaculatus / [Disparidades sexuais determinadas por medições biométricas e hematológicas em Astyanax bimaculatus]

This study aimed to compare the body indexes and hematological characteristics between Astyanax bimaculatus males and females. Four hundred fish were randomly distributed into four polyethylene tanks (100 fish/unit) in a recirculation system and fed four times a day (3% of biomass). After 90 days, ten fish (five ♀ and five ♂) were removed to perform blood tests and to measure weight, height, total length, height/length ratio, condition factor and index determination: vicerosomatic (VSI), hepatosomatic (HSI), and gonadosomatic (GSI). The results showed a higher average weight (g) in females (12.32±0.71) compared to males (6.98±0.75), the same happened to height (cm) = (3.01±0.07) and (2.40±0.05), total length (cm) = (3.01±0.07) and (2.40±0.05), VSI (%) = (11.43±0.81) and (3.55±1.05), HSI (%) = (0.72±0.08) and (0.30±0.04), respectively. Mean corpuscular hemoglobin (pg) was higher in females (3.72±1.20) than in males (2.99±1.51). Regarding the number of thrombocytes (103.µL-1), there was an increase in males (25.71±3.91) compared to females (17.40±6.40).(AU)

O objetivo deste trabalho foi comparar os índices corporais e as características hematológicas entre machos e fêmeas de Astyanax bimaculatus. Quatrocentos peixes foram distribuídos aleatoriamente em quatro caixas de polietileno (100 peixes/unidade), em sistema de recirculação, e alimentados quatro vezes ao dia (3% da biomassa). Após 90 dias, 10 peixes (cinco ♀ e cinco ♂) foram retirados para realização das análises sanguíneas e para mensuração do peso, da altura, do comprimento total, da relação altura/comprimento, do fator de condição e da determinação dos índices: viscerossomático (IVS), hepatossomático (IHS) e gonadossomático (IGS). Os resultados mostraram um maior peso médio (g) nas fêmeas (12,32±0,71) em relação aos machos (6,98±0,75); o mesmo aconteceu para altura (cm) = (3,01± 0,07) e (2,40± 0,05), comprimento total (cm) = (3,01±0,07) e (2,40±0,05), IVS (%) = (11,43±0,81) e (3,55±1,05), IHS (%) = (0,72±0,08) e (0,30±0,04), respectivamente. Hemoglobina corpuscular média (pg) foi maior nas fêmeas (3,72±1,20) que nos machos (2,99±1,51). Em relação ao número de trombócitos (103/µL), houve um aumento nos machos (25,71± 3,91) em relação às fêmeas (17,40±6,40).(AU)

[Mutation signatures in head and neck squamous cell carcinoma : Pathogenesis and therapeutic potential]. / Mutationssignaturen beim Kopf- und Hals-Tumor : Pathogenese und Potenzial für die Therapie.

BACKGROUND: The pathogenesis of head and neck squamous cell carcinoma (HNSCC) is a complex and multistage process which results from the interaction of exogenous and endogenous cellular processes. Each of these processes leaves a characteristic pattern of mutations on the tumor genome, a so-called mutational signature. STATE OF THE ART: The subject of current studies is to decipher specific signatures of mutational processes operating during HNSCC pathogenesis and to address their prognostic value. Computational analysis of genomic sequencing data by The Cancer Genome Atlas (TCGA) revealed mutational signatures 1, 2, 4, 5, 7, and 13 as the main players in HNSCC pathogenesis. Signature 16 was first discovered in human papillomavirus (HPV)-negative oral and oropharyngeal tumors. In many studies, an association of signature 16 with alcohol and tobacco consumption as well as with an unfavorable prognosis was described.

[Gender reassignment surgery from male to female]. / Geschlechtsangleichung von Mann zu Frau.

For many trans*women, the surgical assignment of the male genital into a female is a fundamental part of the transition. Erogenous sensation of the neoclitoris is achieved by meticulous preparation of the penile glans with the neurovascular bundle. Several techniques are available for the formation of a neovagina, the penile inversion technique developed by Burou in the 1950s being the gold standard. With this technique, the inverted penile shaft skin is used as a pedicled flap to line the neovaginal canal. Alternatively, free skin grafts can be used, which serve primarily as a technique for redo procedures. Another technique is the use of intestinal segments to line the vaginal canal. This method is mostly used for redo procedures, but can also be performed primarily if penile skin is too small. Due to the numerous steps involved in the preparation, a wide variety of complications must be expected. Injury to the rectum during dissection of the neovaginal space, with an incidence of 4.5%, represents the greatest challenge. The most common complications are urethra-associated; hereby both a deviation of the urinary stream and strictures of the urethra are possible. The subjective satisfaction of trans*women with the surgical outcome is high and is reported in various studies to be 72-92%. On the basis of validated questionnaires it could also be shown that gender reassignment surgery leads to an increase in the trans*specific quality of life and promotes both subjectively perceived well-being and sexual satisfaction.

Supplementation of Curcuma longa hydrolate improves immunomodulatory response in Nile tilapia reared in a recirculation aquaculture system / [Suplementação de extrato aquoso de Curcuma longa aprimorando a resposta imunomoduladora em tilápias cultivadas em sistema de recirculação em aquicultura]

The objective of this study was to evaluate the effects of dietary supplementation with different doses of Curcuma longa hydrolate on the hematological, immunological and zootechnical parameters of Nile tilapia cultivated in a recirculation system (RAS). Nile tilapia (Oreochromis niloticus) were used, distributed in 16 polyethylene boxes, divided into four treatments: 0.0%; 2.5%; 7.5% and 10.0% of Curcuma longa hydrolate, in quadruplicate. After 45 days of treatment, four fish per experimental unit were anesthetized to remove blood aliquot for hematological and immunological analyzes and dissect the liver to evaluate the hepatosomatic index and final biometry. In the haematological analysis, the fish fed with 2.5% had a higher number of leukocytes, monocytes and lymphocytes than control, while the doses of 7.5% and 10.0% did not differ. Antimicrobial activity showed a significant decrease as the dose of C. longa hydrolate increased. The other hematological, immunological, hepatosomatic index and zootechnical data did not differ between treatments. Thus, supplementation of the hydrolate of Curcuma longa at a dosage of 2.5%, improved and maintained blood-immune homeostasis parameters in these animals, being suggested for further studies.(AU)

O objetivo deste estudo foi avaliar os efeitos da suplementação dietética com diferentes doses de hidrolato de Curcuma longa nos parâmetros hematológicos, imunológicos e zootécnicos da tilápia-do-nilo cultivada em sistema de recirculação. Utilizou-se tilápia- do-nilo (Oreochromis niloticus), distribuída em 16 caixas de polietileno, divididas em quatro tratamentos: 0,0%; 2,5%7,5% e 10,0% de hidrolato de Curcuma longa, em quadruplicata. Após 45 dias de tratamento, quatro peixes por unidade experimental foram anestesiados para remover uma alíquota sanguínea para análises hematológicas e imunológicas, e removeu-se o fígado para avaliar o índice hepatossomático e a biometria final. Na análise hematológica, os peixes alimentados com 2,5% apresentaram maior número de leucócitos, monócitos e linfócitos que no controle, enquanto as doses de 7,5% e 10,0% não diferiram. Por outro lado, a atividade antimicrobiana apresentou uma redução significativa à medida que a dose de hidrolato de C. longa aumentou. Os demais dados hematológicos, imunológicos, hepatossomáticos e zootécnicos não diferiram entre os tratamentos. Assim, a suplementação do hidrolato de Curcuma longa, na dose de 2,5%, melhorou os parâmetros hematoimunológicos e manteve a homeostase nesses animais, sendo sugerida para novos estudos.(AU)

Novel multiparameter correlates of Coxiella burnetii infection and vaccination identified by longitudinal deep immune profiling.

Q-fever is a flu-like illness caused by Coxiella burnetii (Cb), a highly infectious intracellular bacterium. There is an unmet need for a safe and effective vaccine for Q-fever. Correlates of immune protection to Cb infection are limited. We proposed that analysis by longitudinal high dimensional immune (HDI) profiling using mass cytometry combined with other measures of vaccination and protection could be used to identify novel correlates of effective vaccination and control of Cb infection. Using a vaccine-challenge model in HLA-DR transgenic mice, we demonstrated significant alterations in circulating T-cell and innate immune populations that distinguished vaccinated from naïve mice within 10 days, and persisted until at least 35 days post-vaccination. Following challenge, vaccinated mice exhibited reduced bacterial burden and splenomegaly, along with distinct effector T-cell and monocyte profiles. Correlation of HDI data to serological and pathological measurements was performed. Our data indicate a Th1-biased response to Cb, consistent with previous reports, and identify Ly6C, CD73, and T-bet expression in T-cell, NK-cell, and monocytic populations as distinguishing features between vaccinated and naïve mice. This study refines the understanding of the integrated immune response to Cb vaccine and challenge, which can inform the assessment of candidate vaccines for Cb.

MK-8719, a Novel and Selective O-GlcNAcase Inhibitor That Reduces the Formation of Pathological Tau and Ameliorates Neurodegeneration in a Mouse Model of Tauopathy.

Deposition of hyperphosphorylated and aggregated tau protein in the central nervous system is characteristic of Alzheimer disease and other tauopathies. Tau is subject to O-linked N-acetylglucosamine (O-GlcNAc) modification, and O-GlcNAcylation of tau has been shown to influence tau phosphorylation and aggregation. Inhibition of O-GlcNAcase (OGA), the enzyme that removes O-GlcNAc moieties, is a novel strategy to attenuate the formation of pathologic tau. Here we described the in vitro and in vivo pharmacological properties of a novel and selective OGA inhibitor, MK-8719. In vitro, this compound is a potent inhibitor of the human OGA enzyme with comparable activity against the corresponding enzymes from mouse, rat, and dog. In vivo, oral administration of MK-8719 elevates brain and peripheral blood mononuclear cell O-GlcNAc levels in a dose-dependent manner. In addition, positron emission tomography imaging studies demonstrate robust target engagement of MK-8719 in the brains of rats and rTg4510 mice. In the rTg4510 mouse model of human tauopathy, MK-8719 significantly increases brain O-GlcNAc levels and reduces pathologic tau. The reduction in tau pathology in rTg4510 mice is accompanied by attenuation of brain atrophy, including reduction of forebrain volume loss as revealed by volumetric magnetic resonance imaging analysis. These findings suggest that OGA inhibition may reduce tau pathology in tauopathies. However, since hundreds of O-GlcNAcylated proteins may be influenced by OGA inhibition, it will be critical to understand the physiologic and toxicological consequences of chronic O-GlcNAc elevation in vivo. SIGNIFICANCE STATEMENT: MK-8719 is a novel, selective, and potent O-linked N-acetylglucosamine (O-GlcNAc)-ase (OGA) inhibitor that inhibits OGA enzyme activity across multiple species with comparable in vitro potency. In vivo, MK-8719 elevates brain O-GlcNAc levels, reduces pathological tau, and ameliorates brain atrophy in the rTg4510 mouse model of tauopathy. These findings indicate that OGA inhibition may be a promising therapeutic strategy for the treatment of Alzheimer disease and other tauopathies.

Library preparation for next generation sequencing: A review of automation strategies.

Next generation sequencing is in the process of evolving from a technology used for research purposes to one which is applied in clinical diagnostics. Recently introduced high throughput and benchtop instruments offer fully automated sequencing runs at a lower cost per base and faster assay times. In turn, the complex and cumbersome library preparation, starting with isolated nucleic acids and resulting in amplified and barcoded DNA with sequencing adapters, has been identified as a significant bottleneck. Library preparation protocols usually consist of a multistep process and require costly reagents and substantial hands-on-time. Considerable emphasis will need to be placed on standardisation to ensure robustness and reproducibility. This review presents an overview of the current state of automation of library preparation for next generation sequencing. Major challenges associated with library preparation are outlined and different automation strategies are classified according to their functional principle. Pipetting workstations allow high-throughput processing yet offer limited flexibility, whereas microfluidic solutions offer great potential due to miniaturisation and decreased investment costs. For the emerging field of single cell transcriptomics for example, microfluidics enable singularisation of tens of thousands of cells in nanolitre droplets and barcoding of the RNA to assign each nucleic acid sequence to its cell of origin. Finally, two applications, the characterisation of bacterial pathogens and the sequencing within human immunogenetics, are outlined and benefits of automation are discussed.

STIP1 Tissue Expression Is Associated with Survival in Chemotherapy-Treated Bladder Cancer Patients.

To optimize treatment decisions in advanced bladder cancer (BC), we aimed to assess the therapy predictive value of STIP1 with regard to cisplatin therapy. Cisplatin-based chemotherapy represents the standard first-line systemic treatment of advanced bladder cancer. Since novel immunooncologic agents are already available for cisplatin-resistant or ineligible patients, biological markers are needed for the prediction of cisplatin resistance. STIP1 expression was analyzed in paraffin-embedded bladder cancer tissue samples of 98 patients who underwent adjuvant or salvage cisplatin-based chemotherapy by using immunohistochemistry. Furthermore, pre-chemotherapy serum STIP1 concentrations were determined in 48 BC patients by ELISA. Results were correlated with the clinicopathological and follow-up data. Stronger STIP1 nuclear staining was associated with worse OS in both the whole patient group (p = 0.034) and the subgroup of patients who received at least 2 cycles of chemotherapy (p = 0.043). These correlations remained significant also in the multivariable analyses (p = 0.035 and p = 0.040). Stronger STIP1 cytoplasmatic immunostaining correlated with shorter PFS both in the whole cohort (p = 0.045) and in the subgroup of patients who received at least 2 cycles of chemotherapy (p = 0.026). Elevated STIP1 serum levels were associated with older patient's age, but we found no correlation between STIP1 serum levels and patients' outcome. Our results suggest that tissue STIP1 analysis might be used for the prediction of cisplatin-resistance in BC. In contrast, pretreatment STIP1 serum levels showed no predictive value for chemotherapy response and survival.

Review on pneumatic operations in centrifugal microfluidics.

Centrifugal microfluidics allows for miniaturization, automation and parallelization of laboratory workflows. The fact that centrifugal forces are always directed radially outwards has been considered a main drawback for the implementation of complex workflows leading to the requirement of additional actuation forces for pumping, valving and switching. In this work, we review and discuss the combination of centrifugal with pneumatic forces which enables transport of even complex liquids in any direction on centrifugal systems, provides actuation for valving and switching, offers alternatives for mixing and enables accurate and precise metering and aliquoting. In addition, pneumatics can be employed for timing to carry out any of the above listed unit operations in a sequential and cascaded manner. Firstly, different methods to generate pneumatic pressures are discussed. Then, unit operations and applications that employ pneumatics are reviewed. Finally, a tutorial section discusses two examples to provide insight into the design process. The first tutorial explains a comparatively simple implementation of a pneumatic siphon valve and provides a workflow to derive optimum design parameters. The second tutorial discusses cascaded pneumatic operations consisting of temperature change rate actuated valving and subsequent pneumatic pumping. In conclusion, combining pneumatic actuation with centrifugal microfluidics allows for the design of robust fluidic networks with simple fluidic structures that are implemented in a monolithic fashion. No coatings are required and the overall demands on manufacturing are comparatively low. We see the combination of centrifugal forces with pneumatic actuation as a key enabling technology to facilitate compact and robust automation of biochemical analysis.