Investigation of UVC Induced DNA Damage Formation and Photolyase Catalyzed Repair of Cyclobutane Pyrimidine Dimers

Gradual depletion of the ozone layer and consequently, increased ultraviolet (UV) radiation on the Earth's surface induces DNA-lesions inside the genome, thereby causing mutations. Three kinds of photoproducts are mainly formed, namely: cyclobutane pyrimidine dimers (CPD), pyrimidine-(6-4)-pyrimidone dimer [(6-4)PP] and the Dewar valence isomer of (6-4)PP lesion. The formation of these photolesions is a major cause of cell death (aging) and fatal disease like skin cancer. A part of this research was performed to investigate the formation and characterization of DNA-lesions under UVC radiation. Small, fluorescent labeled oligonucleotide hairpins (DNA, RNA and mixed DNA-RNA) were employed to this purpose. The amount of damage was quantified using HPLC. Here, a new method was developed, using ion-exchange SAX-column which works at pH = 13, to measure the lesion formation in a direct way. In order to know which lesions are formed, the irradiated hairpins were enzymatically digested. The lesions were separated by HPLC followed by their characterization using MS/MS fragmentation analysis. Investigation was performed to understand the impact of the neighboring nucleobases on the photo-reactivity of a dTpdT-dinucleotide. For this, hairpins were prepared in various sequential contexts. Analysis of these irradiated hairpins revealed the surprising result that the reactivity is strongly reduced when a dTpdT dinucleotide is locked between two neighboring 2'- deoxyguanosines, strongly implying that genomic DNA will be inhomogeneously damaged and hence mutated under UVC radiation. In order to account for the effect of oligonucleotide conformations (A versus B) on the photolesions formation, DNA, RNA and mixed DNA-RNA hairpins were irradiated. The most surprising observation is that the oligonucleotide hairpins, possessing more A-like conformations were found to be very much resistant to UVC degradation. RNA hairpins containing UpU dinucleotides were found to be fully protected from being damaged. A short, dTpdT-containing DNA stretch, embedded in an A-like RNA environment, was also found to be highly stable under UVC light. In the second part of this study, investigation was performed to assay the activities of CPD-photolyase enzyme isolated from different organisms. A synthetic cis-syn thymidine dimer with an open backbone was incorporated into DNA. CPD-photolyases were found to recognize and split this CPDlesion, via a cycloreversion process, into two 2'-deoxythymidine monomers. In order to profile the activities of photolyases and also to study photolyase activity inside living cell, a sensitive DNA-probe, known as molecular beacon was synthesized. The hairpin probe, which features a loop and stem structure contains the model CPD-lesion in its loop region. The molecular beacon (MB 1) in its closed form, is non-fluorescent due to efficient energy transfer (FRET) from the 5'-FAM to 3'-Dabsyl. The MB 1 was designed to undergo strand break on reaction with photolyase in presence of light, thus separating the FAM and the Dabsyl and causing the fluorescence of FAM to be restored. The activities of the photolyase can be studied by monitoring the fluorescence change. Activities of CPDphotolyase isolated from A. nidulans and A. thaliana were studied using this MBprobe. The sensitivity of this probe was tested with wild-type cell-extract from A. thaliana. The fishing-out of photolyase activity from this wild-type extract was possible. Effort was made to investigate the repair process within a living cell, using laser scanning fluorescence microscopy. The insertion of this artificial, chemically modified DNA-substrate in the cell nucleus was achieved

Adenomatoid tumor of the uterus; report of a rare incidentaloma

Adenomatoid tumor of the uterus is extremely rare and usually an incidental finding in the uteri removed surgically for some other cause as in the present case. The histogenesis of this tumor has been controversial ever since its discovery, although the most favored and proposed is a mesothelial origin. Grossly it is usually mistaken for a leiomyoma. The tumor may have a variety of histomorphologic patterns with rare bizarre appearances, at times leading to an erroneous diagnosis of metastatic adenocarcinoma

Uterine leiomyosarcoma metastasizing to multiple sites: a rare presentation

Uterine leiomyosarcoma is a rare malignant neoplasm which has a dismal outcome especially when associated with widespread metastasis. It shows early metastasis primarily to the lungs, peritoneal cavity, bones, pelvic and para-aortic lymph nodes. We report simultaneous metastasis to the scalp, breast and soft tissue from leiomyosarcoma arising in the uterine broad ligament. Leiomyosarcoma poses a diagnostic difficulty at the metastatic site, especially when the primary site is unknown. Correct tumor typing and separating a primary from a metastatic one is important because of the different treatment modalities for both. Fine Needle Aspiration Cytology (FNAC) has emerged as an indispensable preliminary modality in investigating the metastatic disease and therefore cytomorphological recognition of the tumor presumes a great significance

Spectrum of vulvar lesions: a clinicopathologic study of 170 cases

Background: A wide range lesions may occur in the vulvar region. The clinician is often confronted with the challenge to draw a distinction between normal variants, benign entities and a potentially serious pathology. The aim of the present study is to have an insight into the diverse morphologic spectrum of vulvar lesions.Methods: The present retrospective study was carried out by compiling the data from archival records over a period of eight years from January 2005 to December 2012. The vulvar biopsies/vulvectomy specimens were studied for histomorphological features. The lesions were categorized as non neoplastic, neoplastic and inconclusive; neoplastic ones were further divided into benign, malignant and premalignant.Results: The age of the women ranged from 6 to 80 years (mean 38.2±6.4) with the maximum number of patients between 31 to 40 years of age. Most common clinical presentation was itching and white plaque on the vulva (85 cases; 50%). The commonest site of vulval lesions was labia majora (87 cases, 51.18%). Non neoplastic lesions were more common (n = 94; 55.29%) than the neoplastic lesions (n =50; 29.41%). There were 23 (46%) benign lesions while 27 cases (54%) were malignant or premalignant ones. In 26 cases, no definitive histologic diagnosis could be rendered.Conclusion: Early recognition of vulvar lesions and a prompt biopsy diagnosis for all lesions with suspicious changes is of great significance. The term leukoplakia is imprecise and should be replaced by a precise histological description

Perspectives on physics-based one-dimensional modeling of lung physiology

The need to understand how infection spreads to the deep lung was acutely realized during the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) pandemic. The challenge of modeling virus laden aerosol transport and deposition in the airways, coupled with mucus clearance, and infection kinetics, became evident. This perspective provides a consolidated view of coupled one-dimensional physics-based mathematical models to probe multifaceted aspects of lung physiology. Successes of 1D trumpet models in providing mechanistic insights into lung function and optimalities are reviewed while identifying limitations and future directions. Key non-dimensional numbers defining lung function are reported. The need to quantitatively map various pathologies on a physics-based parameter space of non-dimensional numbers (a virtual disease landscape) is noted with an eye on translating modeling to clinical practice. This could aid in disease diagnosis, get mechanistic insights into pathologies, and determine patient specific treatment plan. 1D modeling could be an important tool in developing novel measurement and analysis platforms that could be deployed at point-of-care

Design, synthesis, and evaluation of 1,4-benzothiazine-3-one containing bisamide derivatives as dual inhibitors of Staphylococcus aureus with plausible application in a urinary catheter

In this study, 1,4-benzothiazine-based bisamide derivatives, a new class of antibacterial agents targeting bacterial peptide deformylase (PDF), were designed and synthesized to combat Staphylococcus aureus infection. Molecular modeling of the designed molecules showed better docking scores compared to the natural product actinonin. Bioactivity assessment identified two derivatives with promising antibacterial activity in vitro. The stability of the most active molecule, 8bE, was assessed using molecular dynamics (MD) simulation. Significantly, compound 8bE could also inhibit the S. aureus biofilm at low concentrations. Furthermore, the capability of the synthesized molecule to inhibit S. aureus biofilm formation on medical devices like urinary catheters is also demonstrated

Organic compounds in PM2.5 emitted from fireplace and woodstove combustion of typical Portuguese wood species

The aim of this study is the further characterisation of PM2.5 emissions from the residential wood combustion of common woods grown in Portugal. This new research extends to eight the number of biomass fuels studied and tries to understand the differences that the burning appliance (fireplace versus woodstove) and the combustion temperature (cold and hot start) have on emissions. Pinus pinaster (Maritime pine), Eucalyptus globulus (eucalypt), Quercus suber (cork oak), Acacia longifolia (Golden wattle), Quercus faginea (Portuguese oak), Olea europea (Olive), Quercus ilex rotundifolia (Holm oak) and briquettes produced from forest biomass waste were used in the combustion tests. Determinations included fine particle emission factors, carbonaceous content (OC and EC) by a thermaleoptical transmission technique and detailed identification and quantification of organic compounds by gas chromatographyemass spectrometry. Fine particle emission factors from the woodstove were lower than those from the fireplace. For both combustion appliances, the OC/EC ratio was higher in “cold start” tests (1.56 0.95 for woodstove and 2.03 1.34 for fireplace). These “cold start” OC/EC values were, respectively, for the woodstove and the fireplace, 51% and 69% higher than those obtained in “hot start” experiments. The chromatographically resolved organics included n-alkanes, n-alkenes, PAHs, n-alkanals, ketones, n-alkanols, terpenoids, triterpenoids, phenolic compounds, phytosterols, alcohols, n-alkanoic acids, n-di-acids, unsaturated acids and alkyl esters of acids. The smoke emission rate and composition varied widely depending on fuel type, burning appliance and combustion temperature

Stochastic finite elements of discretely parameterized random systems on domains with boundary uncertainty

The problem of representing random fields describing the material and boundary properties of the physical system at discrete points of the spatial domain is studied in the context of linear stochastic finite element method. A randomly parameterized diffusion system with a set of independent identically distributed stochastic variables is considered. The discretized parametric fields are interpolated within each element with multidimensional Lagrange polynomials and integrated into the weak formulation. The proposed discretized random-field representation has been utilized to express the random fluctuations of the domain boundary with nodal position coordinates and a set of random variables. The description of the boundary perturbation has been incorporated into the weak stochastic finite element formulation using a stochastic isoparametric mapping of the random domain to a deterministic master domain. A method for obtaining the linear system of equations under the proposed mapping using generic finite element weak formulation and the stochastic spectral Galerkin framework is studied in detail. The treatment presents a unified way of handling the parametric uncertainty and random boundary fluctuations for dynamic systems. The convergence behavior of the proposed methodologies has been demonstrated with numerical examples to establish the validity of the numerical scheme

Intravenous doxycycline, azithromycin, or both for severe scrub typhus

BACKGROUND: The appropriate antibiotic treatment for severe scrub typhus, a neglected but widespread reemerging zoonotic infection, is unclear. METHODS: In this multicenter, double-blind, randomized, controlled trial, we compared the efficacy of intravenous doxycycline, azithromycin, or a combination of both in treating severe scrub typhus. Patients who were 15 years of age or older with severe scrub typhus with at least one organ involvement were enrolled. The patients were assigned to receive a 7-day course of intravenous doxycycline, azithromycin, or both (combination therapy). The primary outcome was a composite of death from any cause at day 28, persistent complications at day 7, and persistent fever at day 5. RESULTS: Among 794 patients (median age, 48 years) who were included in the modified intention-to-treat analysis, complications included those that were respiratory (in 62%), hepatic (in 54%), cardiovascular (in 42%), renal (in 30%), and neurologic (in 20%). The use of combination therapy resulted in a lower incidence of the composite primary outcome than the use of doxycycline (33% and 47%, respectively), for a risk difference of −13.3 percentage points (95% confidence interval [CI], (21.6 to −5.1; P=0.002). The incidence with combination therapy was also lower than that with azithromycin (48%), for a risk difference of −14.8 percentage points (95% CI, −23.1 to −6.5; P<0.001). No significant difference was seen between the azithromycin and doxycycline groups (risk difference, 1.5 percentage points; 95% CI, −7.0 to 10.0; P=0.73). The results in the per-protocol analysis were similar to those in the primary analysis. Adverse events and 28-day mortality were similar in the three groups. CONCLUSIONS: Combination therapy with intravenous doxycycline and azithromycin was a better therapeutic option for the treatment of severe scrub typhus than monotherapy with either drug alone. (Funded by the India Alliance and Wellcome Trust; INTREST Clinical Trials Registry–India number, CTRI/2018/08/015159.