Exploiting pretrained biochemical language models for targeted drug design.

MOTIVATION: The development of novel compounds targeting proteins of interest is one of the most important tasks in the pharmaceutical industry. Deep generative models have been applied to targeted molecular design and have shown promising results. Recently, target-specific molecule generation has been viewed as a translation between the protein language and the chemical language. However, such a model is limited by the availability of interacting protein-ligand pairs. On the other hand, large amounts of unlabelled protein sequences and chemical compounds are available and have been used to train language models that learn useful representations. In this study, we propose exploiting pretrained biochemical language models to initialize (i.e. warm start) targeted molecule generation models. We investigate two warm start strategies: (i) a one-stage strategy where the initialized model is trained on targeted molecule generation and (ii) a two-stage strategy containing a pre-finetuning on molecular generation followed by target-specific training. We also compare two decoding strategies to generate compounds: beam search and sampling. RESULTS: The results show that the warm-started models perform better than a baseline model trained from scratch. The two proposed warm-start strategies achieve similar results to each other with respect to widely used metrics from benchmarks. However, docking evaluation of the generated compounds for a number of novel proteins suggests that the one-stage strategy generalizes better than the two-stage strategy. Additionally, we observe that beam search outperforms sampling in both docking evaluation and benchmark metrics for assessing compound quality. AVAILABILITY AND IMPLEMENTATION: The source code is available at https://github.com/boun-tabi/biochemical-lms-for-drug-design and the materials (i.e., data, models, and outputs) are archived in Zenodo at https://doi.org/10.5281/zenodo.6832145. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Identification of Therapeutic Targets for Medulloblastoma by Tissue-Specific Genome-Scale Metabolic Model.

Medulloblastoma (MB), occurring in the cerebellum, is the most common childhood brain tumor. Because conventional methods decline life quality and endanger children with detrimental side effects, computer models are needed to imitate the characteristics of cancer cells and uncover effective therapeutic targets with minimum toxic effects on healthy cells. In this study, metabolic changes specific to MB were captured by the genome-scale metabolic brain model integrated with transcriptome data. To determine the roles of sphingolipid metabolism in proliferation and metastasis in the cancer cell, 79 reactions were incorporated into the MB model. The pathways employed by MB without a carbon source and the link between metastasis and the Warburg effect were examined in detail. To reveal therapeutic targets for MB, biomass-coupled reactions, the essential genes/gene products, and the antimetabolites, which might deplete the use of metabolites in cells by triggering competitive inhibition, were determined. As a result, interfering with the enzymes associated with fatty acid synthesis (FAs) and the mevalonate pathway in cholesterol synthesis, suppressing cardiolipin production, and tumor-supporting sphingolipid metabolites might be effective therapeutic approaches for MB. Moreover, decreasing the activity of succinate synthesis and GABA-catalyzing enzymes concurrently might be a promising strategy for metastatic MB.

Understanding the adaptive laboratory evolution of multiple stress-resistant yeast strains by genome-scale modeling.

Stress responses triggered by external exposures in adaptive laboratory evolution studies alter the ordinary behavior of cells, and the identification of the differences between the starting and the evolved strains would provide ideal strategies to obtain the desired strains. Metabolic networks are one of the most useful tools to analyze data for this purpose. This study integrates differential expression profiles of multiple Saccharomyces cerevisiae strains that have evolved in eight different stress conditions (ethanol, caffeine, coniferyl aldehyde, iron, nickel, phenylethanol, and silver) and enzyme kinetics into a genome-scale metabolic model of yeast, following a new enhanced method. Flux balance analysis, flux variability analysis, robustness, phenotype phase plane, minimization of metabolic adjustment, survivability, sensitivity analyses, and random sampling are conducted to identify the most common and divergent points within strains. Results were examined both individually and comparatively, and the target reactions, metabolites, and enzymes were identified. Our results showed that the models reconstructed by our methodology were able to simulate experimental conditions where efficient protein allocation was the main goal for survival under stressful conditions, and most of the metabolic changes in the adaptation process mainly arose from the differences in the metabolic reactions of energy maintenance (through coenzyme-A and FAD utilization), cell division (folate requirement of DNA synthesis), and cell wall formation (through sterol and ergosterol biosynthesis).

Cell trapping microfluidic chip made of Cyclo olefin polymer enabling two concurrent cell biology experiments with long term durability.

Cyclo Olefin Polymer (COP) based microbioreactors on a microfluidic chip were produced in house by hot-embossing and thermo-compression bonding methods. The chip allows two different experiments to be performed on trapped cells at the same time. On one side of the chip, red fluorescent protein (RFP) tagged nucleolar Nop56 protein was used to track changes in cell cycle as well as protein synthesis within the yeast cells under the application of the anti-tumor agent hydroxyurea (HU). Simultaneously, on the other side of the chip, the response of yeast cells to the drug metformin, mTOR inhibitor, was investigated to reveal the role of TOR signaling in ribosome biogenesis and cell proliferation. The results of 20 h long experiments are captured by taking brightfield and fluorescent microscopy images of the trapped cells every 9 min. The expression of Nop56 protein of ribosome assembly and synthesis was densely observed during G1 phase of cell cycle, and later towards the end of cell cycle the ribosomal protein expression slowed down. Under HU treatment, the morphology of yeast cells changed, but after cessation of HU, the biomass synthesis rate was sustained as monitored by the cell perimeter. Under metformin treatment, the perimeters of single cells were observed to decrease, implying a decrease in biomass growth; however these cells continued their proliferation during and after the drug application. The relation between ribosome biogenesis and cell cycle was successfully investigated on single cell basis, capturing cell-to-cell variations, which cannot be tracked by regular macroscale bioreactors.

VASPIN, ADIPONECTIN AND LEPTIN LEVELS IN TYPE 1 DIABETIC RATS INDUCED BY STREPTOZOTOCIN.

BACKGROUND: Adiponectin, vaspin and leptin are only a few of these numerous adipocytokines. Little is known about the behavior of adipocytokines and how adipose tissue metabolism is affected in this Type 1 DM model. In this study we investigated the serum levels of adiponectin, leptin, vaspin in streptozotocin(STZ) induced diabetic rats. MATERIAL AND METHODS: Twelve Spraque Dawley albino rats were included in the study. The animals were divided into two groups. The first group was diabetic (D) (n: 6) and 60mg / kg STZ was administered intraperitoneally (i.p.) to these rats. The second group was the non-diabetic control (ND) group (n: 6). All the animals were euthanized by cervical dislocation. Quantification of vaspin, Adiponectin, leptin in serum was performed using the ELISA kit. RESULTS: Adiponectin, vaspin levels of diabetic group were found to be statistically lower than of control group (p<0.05). Leptin levels were significantly higher in the diabetic group (P<0.05). CONCLUSION: There is a need for new researches that can explain the relationship between Vaspin, Leptin and Adiponectin and Type 1 diabetes. New studies in this area will open new horizons for the identification of new biomarkers in the diagnosis and treatment of Type 1 diabetes.

A low cost PS based microfluidic platform to investigate cell cycle towards developing a therapeutic strategy for cancer.

Inhibition of DNA damage response pathway in combination with DNA alkylating agents may enhance the selective killing of cancer cells leading to better therapeutic effects. MDM2 binding protein (MTBP) in human has a role in G1 phase (interphase of cell cycle) and its overexpression leads to breast and ovarian cancers. Sld7 is an uncharacterized protein in budding yeast and a potential functional homologue of MTBP. To investigate the role of Sld7 as a therapeutic target, the behavior of the wild-type cells and sld7∆ mutants were monitored in 0.5 nL microbioreactors. The brightfield microscopy images were used to analyze the change in the cell size and to determine the durations of G1 and S/G2/M phases of wild type cells and mutants. With the administration of the alkylating agent, the cell size decreased and the duration of cell cycle increased. The replacement of the medium with the fresh one enabled the cells to repair their DNA. The application of calorie restriction together with DNA alkylating agent to mutant cells resulted in smaller cell size and longer G1 phase compared to those in control environment. For therapeutic purposes, the potential of MTBP in humans or Sld7 in yeast as a drug target deserves further exploration. The fabrication simplicity, robustness and low-cost of this microfluidic bioreactor made of polystyrene allowed us to perform yeast culturing experiments and show a potential for further cell culturing studies. The device can successfully be used for therapeutic applications including the discovery of new anti-microbial, anti-inflammatory, anti-cancer drugs.

Endocan is markedly overexpressed in severe erectile dysfunction.

This study aimed to measure the serum endocan level of patients with erectile dysfunction (ED) and to investigate the possible association between this and vasculogenic severe ED. We performed a prospective analysis of 86 consecutive patients affected by ED. Patients were divided into severe ED (IIEF-5 score < 7) and mild or moderate ED (IIEF-5 score > 7). A strong negative correlation was found between serum endocan levels and peak systolic velocity (p < .001 and r = -.665) in men with severe ED. Univariate logistic regression analysis demonstrated that tobacco consumption (p < .05), serum total 25-hydroxyvitamin D (p < .01), serum endocan levels (p < .01), peak systolic velocity (p < .01), hypertension (p < .001), dyslipidaemia (p < .001), metabolic syndrome (p = .026) and a history of a cardiovascular event (p < .001) significantly increase the risk of severe ED. In the multivariate logistic regression model, we also found that age, hypertension, metabolic syndrome, cardiovascular events and higher serum endocan levels were independently associated with severe ED. Circulating endocan may be used in daily practice as a new marker that correlates with cardiovascular risks and the severity of ED disease.

Fabrication of cyclo olefin polymer microfluidic devices for trapping and culturing of yeast cells.

A microfluidic platform is designed and fabricated to investigate the role of uncharacterized YOR060C (Sld7) protein in aging in yeast cells for the first time. Saccharomyces cerevisiae yeast cells are trapped in the series of C-shaped regions (0.5 nL) of COP (cyclo olefin polymer), PMMA (poly methylmethacrylate), or PS (polystyrene) microbioreactors. The devices are fabricated using hot embossing and thermo-compression bonding methods. Photolithography and electrochemical etching are used to form the steel mold needed for hot embossing. The cell cycle processes are investigated by monitoring green fluorescent protein (GFP) tagged Sld7 expressions under normal as well as calorie restricted conditions. The cells are loaded at 1 µL/min flowrate and trapped successfully within each chamber. The medium is continuously fed at 0.1 µL/min throughout the experiments. Fluorescent signals of the low abundant Sld7 proteins could be distinguished only on COP devices. The background fluorescence of COP is found 1.22 and 7.24 times lower than that of PMMA, and PS, respectively. Hence, experiments are continued with COP, and lasted for more than 40 h without any contamination. The doubling time of the yeast cells are found as 72 min and 150 min, and the growth rates as 9.63 × 10-3 min-1 and 4.62 × 10-3 min-1, in 2% glucose containing YPD and YNB medium, respectively. The product concentration (Sld7p:GFP) increased in accordance with cell growth. The dual role of Sld7 protein in both cell cycle and chronological aging needs to be further investigated following the preliminary experimental results.

Antifibrogenic role of valproic acid in streptozotocin induced diabetic rat penis.

We investigated the therapeutic effects of valproic acid (VPA) on erectile dysfunction and reducing penile fibrosis in streptozocin (STZ)-induced diabetic rats. Eighteen male rats were divided into three experimental groups (Control, STZ-DM, STZ-DM plus VPA) and diabetes was induced by transperitoneal single dose STZ. Eight weeks after, VPA and placebo treatments were given according to groups for 15 days. All rats were anesthetised for the measurement of in vivo erectile response to cavernous nerve stimulation. Afterward penes were evaluated histologically in terms of immune labelling scores of endothelial nitric oxide synthase (eNOS), vascular endothelial growth factor (VEGF) and transforming growth factor-ß1 (TGF-ß1). Slides were also evaluated in terms of collagen/smooth muscle ratio and penile apoptosis. After the treatment with VPA, erectile responses were found as improved when compared with STZ-DM rats but not statistically meaningful. eNOS and VEGF immune expressions diminished in penile corpora of STZ-DM rats and improved with VPA treatment. VPA led to decrease in TGF-ß1 expression and collagen content of diabetic rats' penes. Penile apoptosis was not diminished with VPA. In conclusion, VPA treatment seems to be effective for reducing penile fibrosis in diabetic rats and more prolonged treatment period may enhance erectile functions.

Transcriptional remodeling in response to transfer upon carbon-limited or metformin-supplemented media in S. cerevisiae and its effect on chronological life span.

One of the factors affecting chronological life span (CLS) in budding yeast is nutrient, especially carbon limitation. Aside from metabolites in the growth medium such as glucose, amino acids, and acetic acid, many pharmaceuticals have also been proven to alter CLS. Besides their impact on life span, these drugs are also prospective chemicals to treat the age-associated diseases, so the identification of their action mechanism and their potential side effects is of crucial importance. In this study, the effects of caloric restriction and metformin, a dietary mimetic pharmaceutical, on yeast CLS are compared. Saccharomyces cerevisiae cells grown in synthetic dextrose complete (SDC) up to mid-exponential phase were either treated with metformin or were subjected to glucose limitation. The impacts of these perturbations were analyzed via transcriptomics, and the common (stimulation of glucose uptake, induction of mitochondrial maintenance, and reduction of protein translation) and divergent (stimulation of aerobic respiration and reprogramming of respiratory electron transport chain (ETC)) cellular responses specific to each treatment were determined. These results revealed that both glucose limitation and metformin treatment stimulate CLS extension and involve the mitochondrial function, probably by creating an efficient mitochondria-to-nucleus signaling of either aerobic respiration or ETC signaling stimulation, respectively.

PHISTO: pathogen-host interaction search tool.

SUMMARY: Knowledge of pathogen-host protein interactions is required to better understand infection mechanisms. The pathogen-host interaction search tool (PHISTO) is a web-accessible platform that provides relevant information about pathogen-host interactions (PHIs). It enables access to the most up-to-date PHI data for all pathogen types for which experimentally verified protein interactions with human are available. The platform also offers integrated tools for visualization of PHI networks, graph-theoretical analysis of targeted human proteins, BLAST search and text mining for detecting missing experimental methods. PHISTO will facilitate PHI studies that provide potential therapeutic targets for infectious diseases. AVAILABILITY: http://www.phisto.org. CONTACT: saliha.durmus@boun.edu.tr SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Uncovering the Effect of SARS-CoV-2 on Liver Metabolism via Genome-Scale Metabolic Modeling for Reprogramming and Therapeutic Strategies.

Genome-scale metabolic models (GEMs) are promising computational tools that contribute to elucidating host-virus interactions at the system level and developing therapeutic strategies against viral infection. In this study, the effect of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) on liver metabolism was investigated using integrated GEMs of human hepatocytes and SARS-CoV-2. They were generated for uninfected and infected hepatocytes using transcriptome data. Reporter metabolite analysis resulted in significant transcriptional changes around several metabolites involved in xenobiotics, drugs, arachidonic acid, and leukotriene metabolisms due to SARS-CoV-2 infection. Flux balance analysis and minimization of metabolic adjustment approaches unraveled possible virus-induced hepatocellular reprogramming in fatty acid, glycerophospholipid, sphingolipid cholesterol, and folate metabolisms, bile acid biosynthesis, and carnitine shuttle among others. Reaction knockout analysis provided critical reactions in glycolysis, oxidative phosphorylation, purine metabolism, and reactive oxygen species detoxification subsystems. Computational analysis also showed that administration of dopamine, glucosamine, D-xylose, cysteine, and (R)-3-hydroxybutanoate contributes to alleviating viral infection. In essence, the reconstructed host-virus GEM helps us understand metabolic programming and develop therapeutic strategies to battle SARS-CoV-2.

Applications of 3D Bioprinting Technology to Brain Cells and Brain Tumor Models: Special Emphasis to Glioblastoma.

Primary brain tumor is one of the most fatal diseases. The most malignant type among them, glioblastoma (GBM), has low survival rates. Standard treatments reduce the life quality of patients due to serious side effects. Tumor aggressiveness and the unique structure of the brain render the removal of tumors and the development of new therapies challenging. To elucidate the characteristics of brain tumors and examine their response to drugs, realistic systems that mimic the tumor environment and cellular crosstalk are desperately needed. In the past decade, 3D GBM models have been presented as excellent platforms as they allowed the investigation of the phenotypes of GBM and testing innovative therapeutic strategies. In that scope, 3D bioprinting technology offers utilities such as fabricating realistic 3D bioprinted structures in a layer-by-layer manner and precisely controlled deposition of materials and cells, and they can be integrated with other technologies like the microfluidics approach. This Review covers studies that investigated 3D bioprinted brain tumor models, especially GBM using 3D bioprinting techniques and essential parameters that affect the result and quality of the study like frequently used cells, the type and physical characteristics of hydrogel, bioprinting conditions, cross-linking methods, and characterization techniques.

Exploring data-driven chemical SMILES tokenization approaches to identify key protein-ligand binding moieties.

Machine learning models have found numerous successful applications in computational drug discovery. A large body of these models represents molecules as sequences since molecular sequences are easily available, simple, and informative. The sequence-based models often segment molecular sequences into pieces called chemical words, analogous to the words that make up sentences in human languages, and then apply advanced natural language processing techniques for tasks such as de novo drug design, property prediction, and binding affinity prediction. However, the chemical characteristics and significance of these building blocks, chemical words, remain unexplored. To address this gap, we employ data-driven SMILES tokenization techniques such as Byte Pair Encoding, WordPiece, and Unigram to identify chemical words and compare the resulting vocabularies. To understand the chemical significance of these words, we build a language-inspired pipeline that treats high affinity ligands of protein targets as documents and selects key chemical words making up those ligands based on tf-idf weighting. The experiments on multiple protein-ligand affinity datasets show that despite differences in words, lengths, and validity among the vocabularies generated by different subword tokenization algorithms, the identified key chemical words exhibit similarity. Further, we conduct case studies on a number of target to analyze the impact of key chemical words on binding. We find that these key chemical words are specific to protein targets and correspond to known pharmacophores and functional groups. Our approach elucidates chemical properties of the words identified by machine learning models and can be used in drug discovery studies to determine significant chemical moieties.

Neuroinflammation, Energy and Sphingolipid Metabolism Biomarkers Are Revealed by Metabolic Modeling of Autistic Brains.

Autism spectrum disorders (ASD) are a heterogeneous group of neurodevelopmental disorders generally characterized by repetitive behaviors and difficulties in communication and social behavior. Despite its heterogeneous nature, several metabolic dysregulations are prevalent in individuals with ASD. This work aims to understand ASD brain metabolism by constructing an ASD-specific prefrontal cortex genome-scale metabolic model (GEM) using transcriptomics data to decipher novel neuroinflammatory biomarkers. The healthy and ASD-specific models are compared via uniform sampling to identify ASD-exclusive metabolic features. Noticeably, the results of our simulations and those found in the literature are comparable, supporting the accuracy of our reconstructed ASD model. We identified that several oxidative stress, mitochondrial dysfunction, and inflammatory markers are elevated in ASD. While oxidative phosphorylation fluxes were similar for healthy and ASD-specific models, and the fluxes through the pathway were nearly undisturbed, the tricarboxylic acid (TCA) fluxes indicated disruptions in the pathway. Similarly, the secretions of mitochondrial dysfunction markers such as pyruvate are found to be higher, as well as the activities of oxidative stress marker enzymes like alanine and aspartate aminotransferases (ALT and AST) and glutathione-disulfide reductase (GSR). We also detected abnormalities in the sphingolipid metabolism, which has been implicated in many inflammatory and immune processes, but its relationship with ASD has not been thoroughly explored in the existing literature. We suggest that important sphingolipid metabolites, such as sphingosine-1-phosphate (S1P), ceramide, and glucosylceramide, may be promising biomarkers for the diagnosis of ASD and provide an opportunity for the adoption of early intervention for young children.

Understanding the ADHD-Gut Axis by Metabolic Network Analysis.

Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder diagnosed with hyperactivity, impulsivity, and a lack of attention inconsistent with the patient's development level. The fact that people with ADHD frequently experience gastrointestinal (GI) dysfunction highlights the possibility that the gut microbiome may play a role in this condition. The proposed research aims to determine a biomarker for ADHD by reconstructing a model of the gut-microbial community. Genome-scale metabolic models (GEM) considering the relationship between gene-protein-reaction associations are used to simulate metabolic activities in organisms of gut. The production rates of dopamine and serotonin precursors and the key short chain fatty acids which affect the health status are determined under three diets (Western, Atkins', Vegan) and compared with those of healthy people. Elasticities are calculated to understand the sensitivity of exchange fluxes to changes in diet and bacterial abundance at the species level. The presence of Bacillota (genus Coprococcus and Subdoligranulum), Actinobacteria (genus Collinsella), Bacteroidetes (genus Bacteroides), and Bacteroidota (genus Alistipes) may be possible gut microbiota indicators of ADHD. This type of modeling approach taking microbial genome-environment interactions into account helps us understand the gastrointestinal mechanisms behind ADHD, and establish a path to improve the quality of life of ADHD patients.

Circadian clock crosstalks with autism.

BACKGROUND: The mechanism underlying autism spectrum disorder (ASD) remains incompletely understood, but researchers have identified over a thousand genes involved in complex interactions within the brain, nervous, and immune systems, particularly during the mechanism of brain development. Various contributory environmental effects including circadian rhythm have also been studied in ASD. Thus, capturing the global picture of the ASD-clock network in combined form is critical. METHODS: We reconstructed the protein-protein interaction network of ASD and circadian rhythm to understand the connection between autism and the circadian clock. A graph theoretical study is undertaken to evaluate whether the network attributes are biologically realistic. The gene ontology enrichment analyses provide information about the most important biological processes. RESULTS: This study takes a fresh look at metabolic mechanisms and the identification of potential key proteins/pathways (ribosome biogenesis, oxidative stress, insulin/IGF pathway, Wnt pathway, and mTOR pathway), as well as the effects of specific conditions (such as maternal stress or disruption of circadian rhythm) on the development of ASD due to environmental factors. CONCLUSION: Understanding the relationship between circadian rhythm and ASD provides insight into the involvement of these essential pathways in the pathogenesis/etiology of ASD, as well as potential early intervention options and chronotherapeutic strategies for treating or preventing the neurodevelopmental disorder.

Comprehensive genome-scale metabolic model of the human pathogen Cryptococcus neoformans: A platform for understanding pathogen metabolism and identifying new drug targets.

Introduction: The fungal priority pathogen Cryptococcus neoformans causes cryptococcal meningoencephalitis in immunocompromised individuals and leads to hundreds of thousands of deaths per year. The undesirable side effects of existing treatments, the need for long application times to prevent the disease from recurring, the lack of resources for these treatment methods to spread over all continents necessitate the search for new treatment methods. Methods: Genome-scale models have been shown to be valuable in studying the metabolism of many organisms. Here we present the first genome-scale metabolic model for C. neoformans, iCryptococcus. This comprehensive model consists of 1,270 reactions, 1,143 metabolites, 649 genes, and eight compartments. The model was validated, proving accurate when predicting the capability of utilizing different carbon and nitrogen sources and growth rate in comparison to experimental data. Results and Discussion: The compatibility of the in silico Cryptococcus metabolism under infection conditions was assessed. The steroid and amino acid metabolisms found in the essentiality analyses have the potential to be drug targets for the therapeutic strategies to be developed against Cryptococcus species. iCryptococcus model can be applied to explore new targets for antifungal drugs along with essential gene, metabolite and reaction analyses and provides a promising platform for elucidation of pathogen metabolism.

Serum carbohydrate antigen 19-9 levels in patients with unilateral hydronephrosis due to urinary lithiasis.

AIM: We aimed to investigate that the association between serum carbohydrate antigen 19-9 (CA 19-9) levels and unilateral hydronephrosis due to urinary lithiasis in homogeneous groups who had no urinary tract infection and renal impairment. METHODS: A total of 50 patients with ureteral stone enrolled in this study prospectively. The patients were divided into two groups according to degree of hydronephrosis. Patients without hydronephrosis were taken into Group I and who had hydronephrosis were taken into Group II. All patients were underwent treatment of shock wave lithotripsy (SWL). Serum CA 19-9 was measured using immunassay method pre-treatment of SWL. After complete stone clearence and recovery of hydronephrosis in all patients, patiens were re-evaluated and CA 19-9 levels were measured. RESULTS: There were no significant differences for age, gender, body mass index, stone volume, shock wave number, number of SWL seasons, and serum creatinin between the groups (P>0.05). Pre-SWL serum mean Ca 19-9 levels were 14.82±14.64 U/mL. in Group I and 13.89±13.03 U/mL. in Group II (P=0.8686). After complete stone clearance and recovery of hydronephosis with SWL, the mean CA 19-9 values were measured as 14.14±10.67 U/mL. in Group II and there were not found significant change as compared pre-SWL values for Group II (P=0.7334). CONCLUSION: The relationship between CA 19-9 values and hydronephrosis was not found to be statistically meaningful on the contrary to previous reports and it does not appear as a useful parameter to predict of urinary obstruction non-invasively.

Fabrication Protocol for Thermoplastic Microfluidic Devices: Nanoliter Volume Bioreactors for Cell Culturing.

Microfluidic devices consist of microchannels etched or embossed into substrates made of polymer, glass or silicon. Intricate connections of the microchannels to reactors with some smart mechanical structures such as traps or curvatures fulfil the desired functionalities such as mixing, separation, flow control or setting the environment for biochemical reactions. Here, we describe the fabrication methods of a thermoplastic microbioreactor step by step. First, material selection is made, then, production methods are determined with the equipment that can be easily procured in a laboratory. COP with its outstanding characteristics among many polymers was chosen. Two types of microbioreactors, with and without electrodes, are designed with AutoCAD and L-edit softwares. Photolithography and electrochemical wet etching are used for master mold preparation. Thermal evaporator is employed for pure chromium and gold deposition on COP substrate and etchants are used to form the interdigitated electrodes. Once the master mold produced, hot embossing is used to obtain the designed shape on drilled and planarized COP. Cover COP, with or without electrodes, is bonded to the hot embossed COP via thermo-compression and thermoplastic microfluidic device is realized. Tubings are connected to the device and a bridge between the macro and micro world is established. Yeast or mammalian cells labeled or tagged with GFP/RFP on specific gene products are loaded into the microfluidic device, and real time data on cell dimensions and fluorescence intensity are collected using inverted fluorescence microscope, and finally image processing is used to analyze the acquired data.