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1.
Ultrason Sonochem ; 107: 106914, 2024 Jul.
Article in English | MEDLINE | ID: mdl-38788353

ABSTRACT

Ultrasound-assisted extraction (UAE) is an innovative process for recovering valuable substances and compounds from plants and various biomaterials. This technology holds promise for resource recovery while maintaining the quality of the extracted products. The review comprehensively discusses UAE's mechanism, applications, advantages, and limitations, focusing on extracting essential oils (EOs) from diverse terrestrial plant materials. These oils exhibit preservation, flavor enhancement, antimicrobial action, antioxidant effects, and anti-inflammatory benefits due to the diverse range of specific compounds in their composition. Conventional extraction techniques have been traditionally employed, and their limitations have prompted the introduction of novel extraction methods. Therefore, the review emphasizes that the use of UAE, alone or in combination with other cutting-edge technologies, can enhance the extraction of EOs. By promoting resource recovery, reduced energy consumption, and minimal solvent use, UAE paves the way for a more sustainable approach to harnessing the valuable properties of EOs. With its diverse applications in food, pharmaceuticals, and other industries, further research into UAE and its synergies with other cutting-edge technologies is required to unlock its full potential in sustainable resource recovery and product quality preservation.


Subject(s)
Oils, Volatile , Ultrasonic Waves , Oils, Volatile/chemistry , Oils, Volatile/isolation & purification , Chemical Fractionation/methods , Plants/chemistry
2.
Head Neck Pathol ; 18(1): 10, 2024 Feb 23.
Article in English | MEDLINE | ID: mdl-38393488

ABSTRACT

BACKGROUND: Pleomorphic adenoma is a well-known benign salivary gland neoplasm characterized by the presence of varying proportions of three different components, including bi-layered ducts, myoepithelial cells, and admixed within a chondromyxoid/fibrous stroma. METHOD: We report an interesting case of an adult male who presented with bleeding from an extensively degenerated parotid gland mass, concerning for a vascular neoplasm versus primary malignant tumor. Microscopically, majority of the viable tumor exhibited diffuse proliferation of spindle to epithelioid cells, with focal areas depicting cribriform glands, ducts, and scant chondromyxoid stroma. RESULT: Next-generation sequencing (NGS) RNA-based fusion panel analysis identified a gene rearrangement involving the pleomorphic adenoma gene 1 (PLAG1), with a novel, cryptogenic fusion partner known as LINC01606; [LINC01606::PLAG1; inv(8;8)(8q12.1;8q12.1)]. CONCLUSION: To the best of our knowledge, this is the first documented case of a long non-coding RNA (lnc-RNA) serving as a rearrangement partner with the PLAG1 gene. We reviewed the molecular characteristics of this entity and explored the potential role of LINC01606::PLAG1 in the tumorigenesis of pleomorphic adenoma.


Subject(s)
Adenoma, Pleomorphic , Salivary Gland Neoplasms , Adult , Male , Humans , Adenoma, Pleomorphic/genetics , Adenoma, Pleomorphic/pathology , DNA-Binding Proteins/genetics , Salivary Gland Neoplasms/genetics , Salivary Gland Neoplasms/pathology , Gene Rearrangement , RNA
3.
J Pak Med Assoc ; 73(Suppl 4)(4): S52-S55, 2023 Apr.
Article in English | MEDLINE | ID: mdl-37482830

ABSTRACT

Objectives: To examine the C-X-C Motif Chemokine Receptor 1 expression in breast cancer tissues prior to neo-adjuvant chemotherapy, and its relationship to neo-adjuvant chemotherapy effectiveness and other prognostic variables. Method: The prospective study was conducted at Kafrelsheikh University Hospital, Egypt, from November 2018 to March 2021, and comprised patients with recent histopathologically proven breast cancer cases eligible for chemotherapy. Paraffin blocks of tumourspecimens were stained by immunohistochemicalstain using concentrating rabbit anti-human C-X-C Motif Chemokine Receptor 1 polyclonal antibody kits. C-X-C Motif Chemokine Receptor 1 expression was classified into low and high categories. Patients were followed for 2 years for treatment response, disease recurrence and mortality. Data was analysed using SPSS 25. RESULTS: Of the 100 females with mean age 50.2±12.1 years, 52(52%) had their left side affected, while 48(48%) had their rightside affected. There were 52(52%) cases with mean age 49.2±12.9 years having high C-X-C Motif Chemokine Receptor 1 expresssion, while 48(48%) with mean age 51.4±11.2 years had low expression. There was a significant association between high expression and advanced tumour grade, advanced tumourstage, higher frequency of triple negative breast cancer and higher frequency of Ki-67-positive cancers (p<0.05). Patients with high C-X-C Motif Chemokine Receptor 1 expression had significantly lower frequency of complete pathological response when compared with patients with low expression (p<0.001). Patients with high expression had higher frequency of recurrence, shorter disease-free survival, higher mortality and shorter overall survival, but the difference was not significant (p>0.05). Multivariate logistic regression analysis identified triple negative hormonal status (p=0.031) and high baseline C-X-C Motif Chemokine Receptor 1 expression (p<0.001) as significant predictors of complete pathological response. CONCLUSIONS: There was found to be a link between baseline C-X-C Motif Chemokine Receptor 1 expression in breast cancer tissues and pathological response to neoadjuvant therapy in breast cancer patients.


Subject(s)
Breast Neoplasms , Triple Negative Breast Neoplasms , Female , Humans , Adult , Middle Aged , Breast Neoplasms/pathology , Neoadjuvant Therapy , Prospective Studies , Retrospective Studies , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Neoplasm Recurrence, Local , Prognosis , Triple Negative Breast Neoplasms/drug therapy , Triple Negative Breast Neoplasms/pathology , Chemotherapy, Adjuvant , Receptors, Chemokine/therapeutic use , Receptor, ErbB-2/metabolism
4.
J Pak Med Assoc ; 73(Suppl 4)(4): S200-S204, 2023 Apr.
Article in English | MEDLINE | ID: mdl-37482858

ABSTRACT

Objectives: To examine the chemokine receptor type 1 expression in breast cancer tissues before and after neoadjuvant chemotherapy, and its relationship with pathological response to neoadjuvant chemotherapy and other clinical variables. Method: The prospective study was conducted at Kafrelsheikh University Hospital, Egypt, from November 2018 to March 2021, and comprised female patients with new histopathologically proven breast cancer eligible for chemotherapy. Paraffin blocks of tumourspecimens were stained immunohistochemically using concentrated rabbit anti-human chemokine receptor type 1 polyclonal antibody kits. The patients were followed up for treatment response, disease recurrence and mortality. Data was analysed using SPSS 25. RESULTS: Of the 100 patients with mean age 50.2±12.1 years, 40(40%) in group A with mean age 55.1±9.3 showed marked response and 60(60%) in group B with mean age 47.0±12.7 yearsshowed mild/moderate response (p<0.001). Group A patients had significantly lower baseline and post-treatment chemokine receptor type 1 expression compared to group B patients (p<0.05). The change in chemokine receptor type 1 expression was not significantly different (p>0.05). Patients with tumour grade 3 had significantly higher baseline chemokine receptor type 1 expression compared to patients with tumour grade 2. Tumourstage and post-treatment chemokine receptor type 1 expression were also significantly interlinked (p<0.05). Multivariate regression analysisidentified patients'age, baseline chemokine receptor type 1 and post-treatment chemokine receptor type 1 expressions as predictors of treatment response. CONCLUSIONS: There was found to be an association between baseline and post-treatment chemokine receptor type 1 expression in breast cancer tissues and pathological response to neoadjuvant chemo therapy in such patients.


Subject(s)
Clinical Relevance , Neoadjuvant Therapy , Female , Rabbits , Animals , Prospective Studies , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Neoplasm Recurrence, Local , Receptor, ErbB-2/metabolism
5.
Ann Clin Lab Sci ; 53(3): 489-493, 2023 May.
Article in English | MEDLINE | ID: mdl-37437943

ABSTRACT

Hemophagocytic lymphohistiocytosis (HLH) is an aggressive, life-threatening condition with sudden onset of a systemic inflammatory response syndrome. The triggers can be apparently non-specific, and the clinical presentations can be very deceptive during a rapidly deteriorating clinical course. Herein, we report a case of a 49-year-old White/Caucasian male with no known past medical history who presented with multi-organ failure, including liver, kidney, and bone marrow, along with disseminated intravascular coagulation. He had a high probability of HLH. Unfortunately, he died ten days after the initial presentation. At autopsy, the liver was necrotic and immunostains revealed diffuse positivity for HSV-1 & 2. The bone marrow was markedly hypocellular with phagocytes containing intact and fragmented red blood cells. There was also disseminated fungal infection involving almost all tissues. PCR, done on frozen tissue samples, revealed Aspergillus fumigatus. The rapid and fatal course of this patient illustrates the sometimes-aggressive course of HLH and the importance of autopsy examination in revealing the underlying etiology for this patient's death.


Subject(s)
Disseminated Intravascular Coagulation , Lymphohistiocytosis, Hemophagocytic , Humans , Male , Middle Aged , Liver , Autopsy , Disseminated Intravascular Coagulation/etiology , Herpesvirus 2, Human
6.
J Prosthodont ; 32(1): 90-93, 2023 Jan.
Article in English | MEDLINE | ID: mdl-36169659

ABSTRACT

Continuous innovation in digital dental technology offers new prospects for creating a complete virtual environment. The technique described adds a facial approach to the conventional digital workflow by incorporating 3D face scans to cone beam computed tomography and intraoral scans. Using this workflow, clinicians can obtain a complete virtual patient for facially generated diagnostic wax up and plan and implement a predictable implant placement and interim prosthesis. This technique provides a full digital workflow for restoratively-driven computer-aided implant planning, guided surgery, and 3D printing of an interim complete-arch fixed implant-supported prosthesis.


Subject(s)
Dental Implants , Spiral Cone-Beam Computed Tomography , Humans , Computer-Aided Design , Dental Prosthesis, Implant-Supported/methods , Dental Implantation, Endosseous/methods , Cone-Beam Computed Tomography/methods
7.
J Cancer Res Clin Oncol ; 149(9): 5853-5859, 2023 Aug.
Article in English | MEDLINE | ID: mdl-36585984

ABSTRACT

BACKGROUND: This study aims to compare the incidence of cardiac events and to identify its predictors in left breast cancer patients receiving adjuvant radiotherapy using breath-hold technique (DIBH) versus free breathing technique (FB). METHODS: We conducted a retrospective multi-center study of two arms; the free breathing arm included 208 patients who were treated with traditional radiotherapy treatment technique, while DIBH arm included 224 patients who were treated with breath-hold technique using The Varian Real-time Position Management (RPM). We retrospectively reviewed the medical records of the patients from January 2010 to December 2017. RESULTS: The mean dose to the heart and left anterior descending artery were significantly lower in the DIBH arm (2.10 ± 0.39 and 6.16 ± 0.18 Gy) compared with (4.29 ± 0.60 Gy and 12.69 ± 0.93 Gy, respectively) in the FB arm. The incidence of cardiac events was higher in the FB arm than in the DIBH arm, but it was not statically significant. Our analysis revealed that age, diabetes, hypertension, smoking, mean LAD dose, and heart mean dose were significant prognostic factors for the occurrence of cardiac events in the breath-hold arm. Hypertension, smoking, as well as heart mean dose were independent risk factors for the occurrence of cardiac events. CONCLUSION: Use of the DIBH technique resulted in a significant reduction in doses to the heart, LAD and lesser cardiac events incidence compared to free breathing.


Subject(s)
Breast Neoplasms , Cardiovascular Diseases , Hypertension , Humans , Female , Breast Neoplasms/epidemiology , Breast Neoplasms/radiotherapy , Radiotherapy, Adjuvant/adverse effects , Retrospective Studies , Incidence , Radiotherapy Dosage , Organs at Risk/radiation effects , Radiotherapy Planning, Computer-Assisted/methods
8.
Ann Clin Lab Sci ; 53(6): 969-973, 2023 Nov.
Article in English | MEDLINE | ID: mdl-38182153

ABSTRACT

Uterine carcinosarcomas (UCS) are aggressive tumors characterized by their biphasic nature, consisting of high-grade epithelial and mesenchymal elements. One component may predominate over the other. We present the case of a 59-year-old female who initially received a diagnosis of endometrial serous carcinoma and presented one year later with a malignant neoplasm in the lung featuring osteosarcomatous differentiation. Notably, the bone scan did not reveal any evidence of a primary bone tumor. However, additional sampling from the endometrium demonstrated a UCS with an osteosarcomatous component.Upon reviewing existing literature, it has been observed that metastases in carcinosarcoma cases generally arise from the carcinomatous component. Conversely, the sarcomatous component typically spreads locally to areas such as the vagina, cervix, or fallopian tubes. The presented case stands out as a unique instance of an undiagnosed UCS manifesting as metastatic osteosarcoma in the lung. This case underscores the complexity and diverse presentations of UCS and emphasizes the importance of comprehensive evaluation in understanding its clinical manifestations.


Subject(s)
Bone Neoplasms , Carcinosarcoma , Endometrial Neoplasms , Osteosarcoma , Female , Humans , Middle Aged , Osteosarcoma/diagnosis , Carcinosarcoma/diagnosis , Bone Neoplasms/diagnostic imaging , Lung
9.
Am J Orthod Dentofacial Orthop ; 162(3): 403-409, 2022 Sep.
Article in English | MEDLINE | ID: mdl-36049869

ABSTRACT

INTRODUCTION: Presurgical infant orthopedics was introduced as an interceptive approach for treating cleft lip and/or palate (CLP). This study aimed to evaluate the intraoral digital impression technique as a viable alternative to conventional impression in infants with unilateral CLP. METHODS: Trios 3-Shape scanner (3Shape, Copenhagen, Denmark) was used for intraoral scanning of the infants' maxillary arches to provide a direct digital scan (DDS). In addition, conventional impressions of the same patients were taken in a hospital setting, and the resultant stone models were digitized using the same scanner to create an indirect digital scan (IDS). Both scans (DDS and IDS) were exported in stereolithography format, and the resultant stereolithography files were imported into computer-assisted-design software (Exocad DentalCAD; exocad GmbH, Darmstadt, Germany) for 3-dimensional surface model superimposition. Differences between the 2 surfaces were quantified in millimeters and visually displayed by a color map. RESULTS: Three-dimensional surface model superimposition of the DDS and IDS scans showed an excellent agreement between both approaches, in which differences ranged from 0.01 mm to 0.1 mm CONCLUSIONS: Intraoral direct digital impression in infants with unilateral CLP is a safe, accurate, and time-efficient technique, which can be a viable alternative to conventional impression. This will aid in overcoming the challenges and complications that are frequently associated with using conventional impressions in infants with unilateral CLP, thus reducing the burden of care not only on the patients' families but also on the care providers.


Subject(s)
Cleft Lip , Cleft Palate , Cleft Lip/surgery , Cleft Palate/surgery , Computer-Aided Design , Dental Impression Technique , Humans , Imaging, Three-Dimensional/methods , Infant , Models, Dental
10.
Dialogues Health ; 1: 100061, 2022 Dec.
Article in English | MEDLINE | ID: mdl-36785640

ABSTRACT

Introduction: Arbaeen is a religious ceremony held annually with the participation of a large number of pilgrims. During the pandemic of Covid-19, a mass gathering of Arbaeen pilgrims can strengthen the transmission of this disease and put more pressure on the health care system of countries. The aim of the study is to evaluate the risk of Arbaeen ceremony in the Covid-19 pandemic in 2021. Materials and methods: A mixed method was performed to perform the study using qualitative and quantitative methods. A national risk assessment tool was used that consisted of hazard identification and assessment, scenario development, exposure assessment, vulnerability assessment and risk estimation. The national risk assessment tool was completed through a focused group discussion and the opinions of 20 representatives of the health system and the level of risks were estimated. Data were collected, analyzed, and integrated by the research team. Results: Based on the results of risk matrix analysis, factors such as the increase in the number of new cases of COVID-19, the probability of exposure to a new variant of COVID-19, the probability of arrival of a new variant of COVID-19 to the country, the probability of increasing referrals to healthcare facilities (score 16), the probability of getting respiratory diseases, and the environmental contamination (score 12) occupied the red area of the matrix and were found to have the highest risk (unbearable risk), that needs immediate action. Conclusion: Preparing for a mass gathering such as Arbaeen is very important in order to reduce the risk of communicable disease, and the first step in improving preparedness is risk assessment and its continuity in the various stages of the ceremony. Therefore, policy makers and planners of such events should conduct risk assessments with the participation of local and national public health authorities regularly.

11.
Metab Eng Commun ; 12: e00170, 2021 Jun.
Article in English | MEDLINE | ID: mdl-33850714

ABSTRACT

Increasing understanding of metabolic and regulatory networks underlying microbial physiology has enabled creation of progressively more complex synthetic biological systems for biochemical, biomedical, agricultural, and environmental applications. However, despite best efforts, confounding phenotypes still emerge from unforeseen interplay between biological parts, and the design of robust and modular biological systems remains elusive. Such interactions are difficult to predict when designing synthetic systems and may manifest during experimental testing as inefficiencies that need to be overcome. Transforming organisms such as Escherichia coli into microbial factories is achieved via several engineering strategies, used individually or in combination, with the goal of maximizing the production of chosen target compounds. One technique relies on suppressing or overexpressing selected genes; another involves introducing heterologous enzymes into a microbial host. These modifications steer mass flux towards the set of desired metabolites but may create unexpected interactions. In this work, we develop a computational method, termed Metabolic Disruption Workflow (MDFlow), for discovering interactions and network disruptions arising from enzyme promiscuity - the ability of enzymes to act on a wide range of molecules that are structurally similar to their native substrates. We apply MDFlow to two experimentally verified cases where strains with essential genes knocked out are rescued by interactions resulting from overexpression of one or more other genes. We demonstrate how enzyme promiscuity may aid cells in adapting to disruptions of essential metabolic functions. We then apply MDFlow to predict and evaluate a number of putative promiscuous reactions that can interfere with two heterologous pathways designed for 3-hydroxypropionic acid (3-HP) production. Using MDFlow, we can identify putative enzyme promiscuity and the subsequent formation of unintended and undesirable byproducts that are not only disruptive to the host metabolism but also to the intended end-objective of high biosynthetic productivity and yield. As we demonstrate, MDFlow provides an innovative workflow to systematically identify incompatibilities between the native metabolism of the host and its engineered modifications due to enzyme promiscuity.

12.
J Oral Maxillofac Surg ; 78(12): 2129-2137, 2020 Dec.
Article in English | MEDLINE | ID: mdl-32916133

ABSTRACT

PURPOSE: The objective of this study was to determine the relationship between the mandibular third molar tooth (Md3) and the adjacent lingual cortical bone and determine the incidence of lingual cortex perforation by Md3s. PATIENTS AND METHODS: This retrospective study was designed and implemented from 100 cone-beam computed tomographic scans (CBCTs) of patients with age ranging from 18 to 65 years old. The primary outcome was to assess the incidence of mandibular third molars (Md3s) with existing lingual cortex perforation by their roots. Perforation was assessed at the level of root apex and the most lingual portion on the apical half of the root. Other outcome variables included average thickness of covering lingual bone in the nonperforation group, lingual cortex morphology, impaction, and demographics. Descriptive statistics were computed. RESULTS: More than half the radiographs showed lingual cortex perforation at the level of root apex and most lingual portion on the apical one half of the root (51.2% and 52.8%, respectively). The average thickness of the covering lingual bone was 1.25 mm around the root apex and 0.93 mm around the most lingual portion on the apical half of the root. The most common lingual cortex morphology was the undercut shape. There was statistically significant association between the presence of Md3 impaction and perforation at both root levels [(P value < .001, Effect size = 0.378) and (P value < .001, Effect size = 0.445)]. CONCLUSIONS: Perforation of the lingual cortex by Md3s, whether erupted or impacted, was found in >50% of patients as determined by a preoperative CBCT scan. Therefore, the finding of lingual cortex perforation after removal of Md3s is likely to be evidence of a pre-existing condition rather than a result of surgery.


Subject(s)
Mandible , Molar, Third , Adolescent , Adult , Aged , Cone-Beam Computed Tomography , Humans , Incidence , Mandible/diagnostic imaging , Mandible/surgery , Middle Aged , Molar , Molar, Third/diagnostic imaging , Molar, Third/surgery , Retrospective Studies , Tooth Root/diagnostic imaging , Young Adult
13.
Mol Psychiatry ; 25(6): 1245-1259, 2020 06.
Article in English | MEDLINE | ID: mdl-31619760

ABSTRACT

Current antidepressants act principally by blocking monoamine reuptake by high-affinity transporters in the brain. However, these antidepressants show important shortcomings such as slow action onset and limited efficacy in nearly a third of patients with major depression disorder. Here, we report the development of a prodrug targeting organic cation transporters (OCT), atypical monoamine transporters recently implicated in the regulation of mood. Using molecular modeling, we designed a selective OCT2 blocker, which was modified to increase brain penetration. This compound, H2-cyanome, was tested in a rodent model of chronic depression induced by 7-week corticosterone exposure. In male mice, prolonged administration of H2-cyanome induced positive effects on several behaviors mimicking symptoms of depression, including anhedonia, anxiety, social withdrawal, and memory impairment. Importantly, in this validated model, H2-cyanome compared favorably with the classical antidepressant fluoxetine, with a faster action on anhedonia and better anxiolytic effects. Integrated Z-scoring across these depression-like variables revealed a lower depression score for mice treated with H2-cyanome than for mice treated with fluoxetine for 3 weeks. Repeated H2-cyanome administration increased ventral tegmental area dopaminergic neuron firing, which may underlie its rapid action on anhedonia. H2-cyanome, like fluoxetine, also modulated several intracellular signaling pathways previously involved in antidepressant response. Our findings provide proof-of-concept of antidepressant efficacy of an OCT blocker, and a mechanistic framework for the development of new classes of antidepressants and therapeutic alternatives for resistant depression and other psychiatric disturbances such as anxiety.


Subject(s)
Antidepressive Agents/pharmacology , Antidepressive Agents/therapeutic use , Depression/drug therapy , Organic Cation Transport Proteins/antagonists & inhibitors , Anhedonia/drug effects , Animals , Antidepressive Agents/administration & dosage , Antidepressive Agents/pharmacokinetics , Anxiety/drug therapy , Disease Models, Animal , Fluoxetine/therapeutic use , Humans , Male , Memory/drug effects , Mice
14.
Microb Cell Fact ; 18(1): 109, 2019 Jun 13.
Article in English | MEDLINE | ID: mdl-31196115

ABSTRACT

BACKGROUND: Metabolic models are indispensable in guiding cellular engineering and in advancing our understanding of systems biology. As not all enzymatic activities are fully known and/or annotated, metabolic models remain incomplete, resulting in suboptimal computational analysis and leading to unexpected experimental results. We posit that one major source of unaccounted metabolism is promiscuous enzymatic activity. It is now well-accepted that most, if not all, enzymes are promiscuous-i.e., they transform substrates other than their primary substrate. However, there have been no systematic analyses of genome-scale metabolic models to predict putative reactions and/or metabolites that arise from enzyme promiscuity. RESULTS: Our workflow utilizes PROXIMAL-a tool that uses reactant-product transformation patterns from the KEGG database-to predict putative structural modifications due to promiscuous enzymes. Using iML1515 as a model system, we first utilized a computational workflow, referred to as Extended Metabolite Model Annotation (EMMA), to predict promiscuous reactions catalyzed, and metabolites produced, by natively encoded enzymes in Escherichia coli. We predict hundreds of new metabolites that can be used to augment iML1515. We then validated our method by comparing predicted metabolites with the Escherichia coli Metabolome Database (ECMDB). CONCLUSIONS: We utilized EMMA to augment the iML1515 metabolic model to more fully reflect cellular metabolic activity. This workflow uses enzyme promiscuity as basis to predict hundreds of reactions and metabolites that may exist in E. coli but may have not been documented in iML1515 or other databases. We provide detailed analysis of 23 predicted reactions and 16 associated metabolites. Interestingly, nine of these metabolites, which are in ECMDB, have not previously been documented in any other E. coli databases. Four of the predicted reactions provide putative transformations parallel to those already in iML1515. We suggest adding predicted metabolites and reactions to iML1515 to create an extended metabolic model (EMM) for E. coli.


Subject(s)
Escherichia coli Proteins/metabolism , Escherichia coli/enzymology , Databases, Protein , Escherichia coli/genetics , Escherichia coli/metabolism , Escherichia coli Proteins/genetics , Metabolome , Metabolomics , Models, Biological
15.
Biotechnol Bioeng ; 116(6): 1405-1416, 2019 06.
Article in English | MEDLINE | ID: mdl-30802311

ABSTRACT

Current pathway synthesis tools identify possible pathways that can be added to a host to produce the desired target molecule through the exploration of abstract metabolic and reaction network space. However, not many of these tools explore gene-level information required to physically realize the identified synthesis pathways, and none explore enzyme-host compatibility. Developing tools that address this disconnect between abstract reactions/metabolic design space and physical genetic sequence design space will enable expedited experimental efforts that avoid exploring unprofitable synthesis pathways. This work describes a workflow, termed Probabilistic Pathway Assembly with Solubility Confidence Scores (ProPASS), which links synthesis pathway construction with the exploration of the physical design space as imposed by the availability of enzymes with predicted characterized activities within the host. Predicted protein solubility propensity scores are used as a confidence level to quantify the compatibility of each pathway enzyme with the host Escherichia coli (E. coli). This study also presents a database, termed Protein Solubility Database (ProSol DB), which provides solubility confidence scores in E. coli for 240,016 characterized enzymes obtained from UniProtKB/Swiss-Prot. The utility of ProPASS is demonstrated by generating genetic implementations of heterologous synthesis pathways in E. coli that target several commercially useful biomolecules.


Subject(s)
Biosynthetic Pathways , Escherichia coli Proteins/metabolism , Escherichia coli/enzymology , Biocatalysis , Escherichia coli/genetics , Escherichia coli/metabolism , Escherichia coli Proteins/genetics , Industrial Microbiology , Metabolic Engineering , Solubility , Workflow
16.
J Neurochem ; 150(3): 330-340, 2019 08.
Article in English | MEDLINE | ID: mdl-30748001

ABSTRACT

Cre/loxP recombination is a widely used approach to study gene function in vivo, using mice models expressing the Cre recombinase under the control of specific promoters or through viral delivery of Cre-expressing constructs. A profuse literature on transgenic mouse lines points out the deleterious effects of Cre expression in various cell types and tissues, presumably by acting on illegitimate loxP-like sites present in the genome. However, most studies reporting the consequences of Cre-lox gene invalidation often omit adequate controls to exclude the potential toxic effects of Cre, compromising the interpretation of data. In this study, we report the anatomical, neurochemical, and behavioral consequences in mice of adeno-associated virus (AAV)-mediated Cre expression in the dopaminergic nuclei substantia nigra, at commonly used viral titers (3 × 109 genome copies/0.3 µL or 2 × 109 genome copies/0.6 µL). We found that injecting AAV-eGFP-Cre into the SN engendered drastic and reproducible modifications of behavior, with increased basal locomotor activity as well as impaired locomotor response to cocaine compared to AAV-eGFP-injected controls. Cre expression in the SN induced a massive decrease in neuronal populations of both pars compacta and pars reticulata and dopamine depletion in the nigrostriatal pathway. This anatomical injury was associated with typical features of programmed cell death, including an increase in DNA break markers, evidence of apoptosis, and disrupted macroautophagy. These observations underscore the need for careful control of Cre toxicity in the brain and the reassessment of previous studies. In addition, our findings suggest that Cre-mediated ablation may constitute an efficient tool to explore the function of specific cell populations and areas in the brain, and the impact of neurodegeneration in these populations.


Subject(s)
Integrases , Neurons/pathology , Substantia Nigra/metabolism , Substantia Nigra/pathology , Animals , Apoptosis/drug effects , Dependovirus , Dopamine/metabolism , Genetic Vectors , Integrases/administration & dosage , Integrases/genetics , Integrases/toxicity , Locomotion/physiology , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neurons/drug effects , Neurons/metabolism
17.
Brain Struct Funct ; 224(3): 1219-1244, 2019 Apr.
Article in English | MEDLINE | ID: mdl-30656447

ABSTRACT

Hevin, also known as SPARC-like 1, is a member of the secreted protein acidic and rich in cysteine family of matricellular proteins, which has been implicated in neuronal migration and synaptogenesis during development. Unlike previously characterized matricellular proteins, hevin remains strongly expressed in the adult brain in both astrocytes and neurons, but its precise pattern of expression is unknown. The present study provides the first systematic description of hevin mRNA distribution in the adult mouse brain. Using isotopic in situ hybridization, we showed that hevin is strongly expressed in the cortex, hippocampus, basal ganglia complex, diverse thalamic nuclei and brainstem motor nuclei. To identify the cellular phenotype of hevin-expressing cells, we used double fluorescent in situ hybridization in mouse and human adult brains. In the mouse, hevin mRNA was found in the majority of astrocytes but also in specific neuronal populations. Hevin was expressed in almost all parvalbumin-positive projection neurons and local interneurons. In addition, hevin mRNA was found in: (1) subsets of other inhibitory GABAergic neuronal subtypes, including calbindin, cholecystokinin, neuropeptide Y, and somatostatin-positive neurons; (2) subsets of glutamatergic neurons, identified by the expression of the vesicular glutamate transporters VGLUT1 and VGLUT2; and (3) the majority of cholinergic neurons from motor nuclei. Hevin mRNA was absent from all monoaminergic neurons and cholinergic neurons of the ascending pathway. A similar cellular profile of expression was observed in human, with expression of hevin in parvalbumin interneurons and astrocytes in the cortex and caudate nucleus as well as in cortical glutamatergic neurons. Furthermore, hevin transcript was enriched in ribosomes of astrocytes and parvalbumin neurons providing a direct evidence of hevin mRNAs translation in these cell types. This study reveals the unique and complex expression profile of the matricellular protein hevin in the adult brain. This distribution is compatible with a role of hevin in astrocytic-mediated adult synaptic plasticity and in the regulation of network activity mediated by parvalbumin-expressing neurons.


Subject(s)
Astrocytes/metabolism , Brain/cytology , Calcium-Binding Proteins/genetics , Calcium-Binding Proteins/metabolism , Extracellular Matrix Proteins/genetics , Extracellular Matrix Proteins/metabolism , Neurons/metabolism , Parvalbumins/metabolism , Adult , Aged , Aged, 80 and over , Animals , Brain Mapping , Excitatory Amino Acid Transporter 1/metabolism , Female , Humans , Male , Mice , Mice, Inbred C57BL , Middle Aged , Nerve Tissue Proteins/metabolism , Postmortem Changes , RNA, Messenger/metabolism , Vesicular Glutamate Transport Proteins/metabolism , Young Adult
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