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Background: Hepatocellular carcinoma (HCC) occurs in a well-defined high-risk patient population, but better screening tests are needed to improve sensitivity and efficacy. Therefore, we investigated the use of urine circulating tumour DNA (ctDNA) as a screening test., Methods: Candidate markers in urine were selected from HCC and controls. We then enrolled 609 patients from five medical centres to test the selected urine panel. A two-stage model was developed to combine AFP and urine panel as a screening test., Results: Mutated TP53, and methylated RASSF1a, and GSTP1 were selected as the urine panel markers. Serum AFP outperformed the urine panel among all cases of HCC, but the urine panel identified 49% of HCC cases with low AFP < 20 ng/ml. Using the two-stage model, the combined AFP and urine panel identified 148 of the 186 HCC cases (79.6% sensitivity at 90% specificity), which was 30% more than the cases detected with serum AFP alone. It also increased early-stage HCC detection from 62% to 92% (BCLC stage 0), and 40% to 77% (BCLC stage A)., Conclusion: Urine ctDNA has promising diagnostic utility in patients in HCC, especially in those with low AFP and can be used as a potential non-invasive HCC screening test., (© 2022. The Author(s), under exclusive licence to Springer Nature Limited.)

Chordoma is a rare form of cancer that forms from the notochord remnants, and affects the skull and the spine. The standard treatment for a sacrococcygeal chordoma is surgery and radiation. Chordoma has a high rate of recurrence and surgery with radiation treatment can leave patients with surgical site complications, such as wounds, fistulas or sinus tracts. Repeat surgical intervention on an irradiated surgical site increases incidence of complications leading to decreased quality of life, and increased morbidity and mortality. Treatment of wound complications after surgery and high-dose radiation for a chordoma tumour is rarely reported in the literature. Herein, the author describes the case of a chronic sinus tract after surgery and radiation for a sacral chordoma tumour successfully healed by hyperbaric oxygen therapy (HBOT) in conjunction with topical antibiotics.

In the present work, a novel smartphone assisted fiber optic (FO)-Long range surface plasmon resonance (LRSPR) based biosensor is proposed. In the developed biosensor, the inbuilt color sensitive property of the digital camera present in the smartphone is used for the monitoring of blue and red color channel intensities. This will replace the most exploited diffraction gratings or narrow band filters used for analyzing the spectral data in reported smartphone based SPR sensors. The proposed technique helps in improving the sensitivity and reduces the chances of wrong detection. For the first time, SiO 2 nanostructured film is employed as the dielectric sensing layer to excite the Long range surface plasmons (LRSPs) in the dielectric-metal-dielectric configuration. The proposed FO-LRSPR biosensor possess limit of detection of 0.02 mM and sensitivity of 0.9/mM and, for uric acid detection in the 0.1 mM-1 mM concentration range. The novel fabricated sensor which is found to be stable up to 24 weeks can be effectively utilized in health sector and environment monitoring and it possess the ability of point-of-care detection, even in rural and remote areas., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Introduction: Developing nations have a huge burden of tuberculosis (TB), because of which many cases are undetected and diagnosed only on autopsies. The purpose of this study was to find these cases in a series of consecutive autopsies conducted at a tertiary care centers., Materials and Methods: This retrospective study of 7-year duration from 2014 to 2020 consisted of reviewing 767 autopsies. The records of cases where a diagnosis of TB were established on histopathology were noted and evaluated., Results: Of 767 autopsies, TB was the diagnosis on histopathology in 72 cases. Of the 72 cases, 53 cases were undiagnosed antemortem and incidentally detected on autopsy. The study showed a male predominance with a 1.52:1 male-to-female ratio. The most common age group involved was the second decade. The study constitutes 26 cases of isolated pulmonary TB, 23 cases of disseminated TB, three cases of solitary renal TB, and one case of hepatic TB. The liver was the most commonly involved organ along with the lung in cases of disseminated TB, followed by the spleen, kidney, pancreas, intestine, brain, and heart., Discussion: Epidemiology and research on TB may be affected by these hidden cases which are unraveled incidentally on autopsy. These cases must always be reported and brought to the attention of clinicians for nation's better public health., Competing Interests: None

The human body possesses an endogenous regeneration system based on stem cells, which may be found in practically every tissue type. They are classified as embryonic stem cells (ESCs) or nonembryonic stem cells (NESCs). Despite its enormous promise, the use of ESCs is presently limited because of ethical and scientific issues. Stem cells have the potential to improve healthcare by using and boosting the body's inherent regenerative capabilities. Although the stem cells offer an enormous promise for tissue regeneration and repair, much more about their biology, administration, and safety must be studied before they may be employed therapeutically. Stem cells and their derivatives will have enormous medical promise in the future. Current animal and laboratory investigations are looking into the viability of bringing stem cell therapy into clinical practice for regeneration in muscular dystrophy, intervertebral disc degeneration, cerebral infarctions, and transplantation medicine. This article delves into the many aspects at play, as well as current situation and possible issues with stem cell treatment in patient care and management (Fig. 1, Ref. 86). Keywords: stem cells, tissue engineering, regenerative medicine, stem cell application.

Thermoanaerobacter kivui is a thermophilic acetogen that can grow on carbon monoxide as sole carbon and energy source. To identify the gene(s) involved in CO oxidation, the genome sequence was analyzed. Two genes potentially encoding CO dehydrogenases were identified. One, cooS, potentially encodes a monofunctional CO dehydrogenase, whereas another, acsA, potentially encodes the CODH component of the CODH/ACS complex. Both genes were cloned, a His-tag encoding sequence was added, and the proteins were produced from a plasmid in T. kivui. His-AcsA copurified by affinity chromatography with AcsB, the acetyl-CoA synthase of the CO dehydrogenase/acetyl CoA synthase complex. His-CooS copurified with CooF 1 , a small iron-sulfur center containing protein likely involved in electron transport. Both protein complexes had CO:ferredoxin oxidoreductase as well as CO:methyl viologen oxidoreductase activity, but the activity of CooSF 1 was 15-times and 231-times lower, respectively. To underline the importance of CooS, the gene was deleted in the CO-adapted strain. Interestingly, the ∆cooS deletion mutant did not grow on CO anymore. These experiments clearly demonstrated that CooS is essential for growth of T. kivui on CO. This is in line with the hypothesis that CooS is the CO-oxidizing enzyme in cells growing on CO., (© 2021. The Author(s).)

Competing Interests: The authors declare no conflict of interest.

Thermoanaerobacter kivui is an acetogenic model organism that reduces CO 2 with electrons derived from H 2 or CO, or from organic substrates in the Wood-Ljugdahl pathway (WLP). For the calculation of ATP yields, it is necessary to know the electron carriers involved in coupling of the oxidative and reductive parts of metabolism. Analyses of key catabolic oxidoreductases in cell-free extract (CFE) or with purified enzymes revealed the physiological electron carriers involved. The glyceraldehyde-3-phosphate dehydrogenase (GA3P-DH) assayed in CFE was NAD + -specific, NADP + was used with less than 4% and ferredoxin (Fd) was not used. The methylene-THF dehydrogenase was NADP + -specific, NAD + or Fd were not used. A Nfn-type transhydrogenase that catalyzes reduced Fd-dependent reduction of NADP + with NADH as electron donor was also identified in CFE. The electron carriers used by the potential electron-bifurcating hydrogenase (HydABC) could not be unambiguously determined in CFE for technical reasons. Therefore, the enzyme was produced homologously in T. kivui and purified by affinity chromatography. HydABC contained 33.9 ± 4.5 mol Fe/mol of protein and FMN; it reduced NADP + but not NAD + . The methylene-THF reductase (MetFV) was also produced homologously in T. kivui and purified by affinity chromatography. MetFV contained 7.2 ± 0.4 mol Fe/mol of protein and FMN; the complex did neither use NADPH nor NADH as reductant but only reduced Fd. In sum, these analysis allowed us to propose a scheme for entire electron flow and bioenergetics in T. kivui., (© 2021. The Author(s).)

Lateral flow assay (LFA) is a flexible, simple, low-costpoint-of-care platform for rapid detection of disease-specific biomarkers. Importantly, the ability of the assay to capture the circulating bio-molecules has gained significant attention, as it offers a potential minimal invasive system for early disease diagnosis and prognosis. In the present article, we review an innovative concept of LFA-based detection of circulating long non-coding RNAs (lncRNAs), one of the key regulators of fundamental biological processes. In addition, their disease-specific expression pattern and presence in biological fluids at differential levels make them excellent biomarker candidates for cancer detection. Our article also provides an update on the requirements for developing and improving such systems and discusses the key aspects of material selection, operational concepts, principles and conceptual design. We assume that the reviewed points will be helpful to improve the diagnostic applicability of LFA based lncRNA detection in cancer diagnosis., Competing Interests: Declaration of Competing Interest The authors report no declarations of interest., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths worldwide. The beta-catenin gene, CTNNB1, is among the most frequently mutated in HCC tissues. However, mutational analysis of HCC tumors is hampered by the difficulty of obtaining tissue samples using traditional biopsy. Here, we explored the feasibility of detecting tumor-derived CTNNB1 mutations in cell-free DNA (cfDNA) extracted from the urine of HCC patients. Using a short amplicon qPCR assay targeting HCC mutational hotspot CTNNB1 codons 32-37 (exon 3), we detected CTNNB1 mutations in 25% (18/73) of HCC tissues and 24% (15/62) of pre-operative HCC urine samples in two independent cohorts. Among the CTNNB1-mutation-positive patients with available matched pre- and post-operative urine ( n = 13), nine showed apparent elimination ( n = 7) or severalfold reduction ( n = 2) of the mutation in urine following tumor resection. Four of the seven patients with no detectable mutations in postoperative urine remained recurrence-free within five years after surgery. In contrast, all six patients with mutation-positive in post-operative urine recurred, including the two with reduced mutation levels. This is the first report of association between the presence of CTNNB1 mutations in pre- and post-operative urine cfDNA and HCC recurrence with implications for minimum residual disease detection.

CRISPR/Cas9 is a powerful genome editing tool, but its off-target cleavage activity can result in unintended adverse outcomes for therapeutic applications. Here we report the design of a simple tunable CRISPR controller in which a chemically inducible anti-CRISPR protein AcrIIA4 is engineered to disable Cas9 DNA binding upon the addition of trimethoprim. Dose-dependent control over Cas9 editing and dCas9 induction was achieved, which drastically improved the specificity and biosafety of the CRISPR/Cas9 system. We utilized the anti-CRISPR protein AcrIIA4 as a means to interfere with Cas9 DNA binding activity. By fusing AcrIIA4 to a ligand-inducible destabilization domain DHFR(DD), we show significantly reduced off-target activity in mammalian cells. Furthermore, we describe a new inducible promoter system Acr-OFF based on CRISPR controllers, which is regulated by an FDA-approved ligand trimethoprim.

Genome editing critically relies on selective recognition of target sites. However, despite recent progress, the underlying search mechanism of genome-editing proteins is not fully understood in the context of cellular chromatin environments. Here, we use single-molecule imaging in live cells to directly study the behavior of CRISPR/Cas9 and TALEN. Our single-molecule imaging of genome-editing proteins reveals that Cas9 is less efficient in heterochromatin than TALEN because Cas9 becomes encumbered by local searches on non-specific sites in these regions. We find up to a fivefold increase in editing efficiency for TALEN compared to Cas9 in heterochromatin regions. Overall, our results show that Cas9 and TALEN use a combination of 3-D and local searches to identify target sites, and the nanoscopic granularity of local search determines the editing outcomes of the genome-editing proteins. Taken together, our results suggest that TALEN is a more efficient gene-editing tool than Cas9 for applications in heterochromatin.

Extending across multiple length scales, dynamic chromatin structure is linked to transcription through the regulation of genome organization. However, no individual technique can fully elucidate this structure and its relation to molecular function at all length and time scales at both a single-cell level and a population level. Here, we present a multitechnique nanoscale chromatin imaging and analysis (nano-ChIA) platform that consolidates electron tomography of the primary chromatin fiber, optical super-resolution imaging of transcription processes, and label-free nano-sensing of chromatin packing and its dynamics in live cells. Using nano-ChIA, we observed that chromatin is localized into spatially separable packing domains, with an average diameter of around 200 nanometers, sub-megabase genomic size, and an internal fractal structure. The chromatin packing behavior of these domains exhibits a complex bidirectional relationship with active gene transcription. Furthermore, we found that properties of PDs are correlated among progenitor and progeny cells across cell division., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Breast cancer is the most commonly occurring cancer among women which leads to thousands of deaths worldwide. The chances of survival are more if the breast cancer is diagnosed at early stage. At present, mammography, magnetic resonance imaging, ultrasound and tissue biopsies are the main diagnostic techniques available for the detection of breast cancer. However, despite of offering promising results, requirement of expensive setup, skilled supervision, expert analysis, invasive procedure (biopsy) and low capacity of multiplexing are the main limitations of these diagnostic techniques. Due to high cost, these screening tests are out of reach of people belonging to low socioeconomic groups and this poses serious health burden to the society. Recently, biosensor-based diagnostic technology for early detection of various types of cancers and other non-oncological disorders have gained considerable attention because of their several advantageous features over existing diagnostic technologies such as high throughput, noninvasive nature, cost effectiveness, easy interpretable results and capacity for multiplexing. Further, biosensors can be designed for biomarkers which are confined to particular type of cancer. In this review, we have discussed about various genomic, transcriptomic, proteomic and metabolomic biomarkers associated with breast cancer, various biosensors-based diagnostic approaches designed for detection of specific biomarkers associated with breast cancer are also described. Further, this review throws insight on various biomarkers linked with breast cancer which can be effectively exploited to develop new diagnostic technology. The assessment of these biomarkers associated with BC using biosensors in large population are cost-effective, non-invasive and high throughput. They help in risk assessment of disease at very initial stage even in backward areas and also help to lower the disease burden of society and economic cost of treatment for a common man. This review would provide new avenues for the development of biosensor based diagnostic technology for the detection of biomarkers associated with breast cancer., (Copyright © 2020 Elsevier Inc. All rights reserved.)

A novel, highly sensitive and low cost Long Range Surface Plasmon Resonance (LRSPR) biosensor for detecting uric acid, as a model analyte, has been developed in this work. Silicon dioxide (SiO 2 ) having low and tunable refractive index has been chosen as the dielectric layer for the excitation of LRSP modes replacing the most explored Cytop and Teflon polymers. The prepared LRSPR based uric acid bio-sensor gives good response characteristics with a high sensitivity of about 21.6°/mM and low limit of detection (LOD) of 0.02 mM. The fabricated LRSPR sensor was also evaluated to detect uric acid in real serum samples. The results yield a great scope to promote the development of robust, efficient and highly selective LRSPR based biosensors with SiO 2 as tunable dielectric layer., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Acetogenic bacteria have gained much attraction in recent years as they can produce different biofuels and biochemicals from H 2 plus CO 2 or even CO alone, therefore opening a promising alternative route for the production of biofuels from renewable sources compared to existing sugar-based routes. However, CO metabolism still raises questions concerning the biochemistry and bioenergetics in many acetogens. In this study, we focused on the two acetogenic bacteria Acetobacterium woodii and Thermoanaerobacter kivui which, so far, are the only identified acetogens harbouring a H 2 -dependent CO 2 reductase and furthermore belong to different classes of 'Rnf'- and 'Ech-acetogens'. Both strains catalysed the conversion of CO into the bulk chemical acetate and formate. Formate production was stimulated by uncoupling the energy metabolism from the Wood-Ljungdahl pathway, and specific rates of 1.44 and 1.34 mmol g -1  h -1 for A. woodii ∆rnf and T. kivui wild type were reached. The demonstrated CO-based formate production rates are, to the best of our knowledge, among the highest rates ever reported. Using mutants of ∆hdcr, ∆cooS, ∆hydBA, ∆rnf and ∆ech2 with deficiencies in key enzyme activities of the central metabolism enabled us to postulate two different CO utilization pathways in these two model organisms., (© 2020 The Authors. Microbial Biotechnology published by Society for Applied Microbiology and John Wiley & Sons Ltd.)

Research on development of novel metal based anti-cancer agents continues with its popularity among bioinorganic community. Benzothiazole, an important heterocyclic pharmacophore, was chosen as a valuable and useful scaffold for the synthesis of novel copper(II) complexes. Three new copper(II) complexes obtained from the synthesis of newly synthesized benzothiazole based N-(benzo[d]thiazol-2-ylmethyl)-N-methyl-2-(pyridin-2-yl)ethan-1-amine (btzpy) ligand with CuCl 2 [Cu(btzpy)Cl 2 ] (1), Cu(NCS) 2 [Cu(btzpy)(NCS) 2 ] (2), and Cu(NO 3 ) 2 [Cu(btzpy)(NO 3 )(H 2 O)]NO 3 (3) were isolated and characterized by physical and spectroscopic measurements, including single-crystal X-ray structures. The interaction of complexes 1 and 3 with calf thymus (CT)-DNA was investigated using ethidium bromide fluorescence quenching assay and weak intercalation with K SV values of 9.8 × 10 2  M -1 and 8.2 × 10 2  M -1 , respectively was observed. All three complexes have shown DNA cleavage of supercoiled plasmid DNA forming single nicked and double nicked forms in the presence of external reducing agents like 3-mercaptopropionic acid (3-MPA) and ascorbic acid. The water-soluble complexes 1 and 3 also show prominent hydrolytic DNA cleavage. From the DPPH (2,2-diphenyl-1-picrylhydrazyl) radical scavenging assay, it was observed that complex 2 also exhibits good antioxidant properties. The cytotoxicity of complexes 1-3 was tested against the lung cancer cell line (A549) and complex 2 with -NCS moiety shows maximum activity in the micromolar range. A rationale for the observed activity is proposed in light of the other properties of these molecules., Competing Interests: Declaration of competing interest There are no competing financial interests to declare., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Here, we report a rapid CRISPR-Cas9-mediated gene knock-in strategy that uses Cas9 ribonucleoprotein and 5'-modified double-stranded DNA donors with 50-base-pair homology arms and achieved unprecedented 65/40% knock-in rates for 0.7/2.5 kilobase inserts, respectively, in human embryonic kidney 293T cells. The identified 5'-end modification led to up to a fivefold increase in gene knock-in rates at various genomic loci in human cancer and stem cells.

Hepatocellular carcinoma (HCC) is the world's second leading cause of cancer death; 82.4% of patients die within 5 years. This grim prognosis is the consequence of a lack of effective early detection tools, limited treatment options, and the high frequency of HCC recurrence. Advances in the field of liquid biopsy hold great promise in improving early detection of HCC, advancing patient prognosis, and ultimately increasing the survival rate. In an effort to address the current challenges of HCC screening and management, several studies have identified and evaluated liver-cancer-associated molecular signatures such as genetic alterations, methylation, and noncoding RNA expression in the form of circulating biomarkers in body fluids and circulating tumor cells of HCC patients. In this review, we summarize the recent progress in HCC liquid biopsy, organized by the intended clinical application of the reported study., (Copyright © 2018. Published by Elsevier Inc.)

Background: Detection of human hepatitis B virus (HBV) DNA in the urine of patients with chronic hepatitis B infection (CHB) has been reported previously, suggesting urine could provide a potential route of horizontal HBV transmission. However, it is not clear whether the HBV DNA detected in urine is indeed full-length, infectious viral DNA. The aim of this study is to assess the potential infectivity of urine from patients with CHB and to correlate HBV DNA detection in urine with clinical parameters, such as serum viral load and HBeAg status., Methods: Urine from 60 CHB patients with serum viral loads ranging from undetectable to 10 8  IU/mL were analyzed for HBV DNA and serum immune markers. HBV DNA was detected from total urine DNA and size-fractionated urine DNA (separated into ≤1 kb and > 1 kb fractions) by PCR analysis of six regions of the HBV genome., Results: Twenty-seven of 59 (45.7%) patients with HBV serum viral load (≥20 IU/mL) contained at least 20 copies per mL of fragmented HBV DNA in urine detected in at least 1 of the 6 PCR assay regions. Only one patient contained HBV DNA detected by all six regions, and was found to have evidence of blood in the urine. Sixteen of 25 urine samples with high viral load (> 10 5  IU/mL) and 11 of 34 urine samples with low viral load (< 10 5  IU/mL) contained detectable HBV DNA. Twelve of 27 (44.44%) patients with detectable HBV DNA in urine were HBeAg positive, and only 5 of these HBeAg positive patients were in the group of 33 (15.15%) patients with no detectable HBV DNA in urine. By Fishers' exact test, HBV DNA in urine is significantly associated with high serum viral load (P = 0.0197) and HBeAg (P = 0.0203)., Conclusions: We conclude that urine from CHB patients with healthy kidney function should not contain full-length HBV DNA, and therefore should not be infectious.

Reported herein the binding affinity between Human Serum Albumin and the DNA binding and cleavage activity of three copper(II) complexes, [Cu(phen)(o-van)ClO 4 ] (1), [Cu(phen)(gly)]ClO 4 (2) and [Cu(L 1 ) 2 (H 2 O) 2 ] (3) wherein 1 and 2 are synthesized with 1,10-phenanthroline (phen) and co-ligands (o-van: o-vanillin; gly: glycine) and 3 with a ligand 2-hydroxy-3-methoxybenzylidene-4H-1,2,4-triazol-4-amine (H 1 L 1 ). Complex 2 crystallizes in monoclinic (P21/n) space group shows square pyramidal geometry. The complex 3 crystallizes in monoclinic (P21/a) space group. All the three complexes exhibit binding affinity towards the transport protein Human Serum albumin (HSA). Quantitative evaluation of the thermodynamics of interaction and the results obtained from fluorescence spectroscopy suggest that metal coordinated glycynate, o-vanillin and perchlorate groups have a major role to play in the binding process, the latter two being stronger in the binding of complex 1. The coordinated water in complex 3 also plays an important role in the binding, which makes binding of complex 3 with HSA stronger than that of complex 2. Experimental results indicate that the binding affinity of the complexes towards CT-DNA is in the order 1>3>2 implying that complex 1 binds stronger than complex 3 and 2.The DNA cleaving activity of all the three complexes was explored in the presence of reactive oxygen compound, H 2 O 2 . All the three complexes have primarily shown the DNA cleaving activity., (Copyright © 2017. Published by Elsevier B.V.)

Early detection of hepatocellular carcinoma (HCC) is critical for the effective treatment. Alpha fetoprotein (AFP) serum level is currently used for HCC screening, but the cutoff of the AFP test has limited sensitivity (~50%), indicating a high false negative rate. We have successfully demonstrated that cancer derived DNA biomarkers can be detected in urine of patients with cancer and can be used for the early detection of cancer (Jain et al., 2015; Lin et al., 2011; Song et al., 2012; Su, Lin, Song, & Jain, 2014; Su, Wang, Norton, Brenner, & Block, 2008). By combining urine biomarkers (uBMK) values and serum AFP (sAFP) level, a new classification model has been proposed for more efficient HCC screening. Several criterions have been discussed to optimal the cutoff for uBMK score and sAFP score. A joint distribution of sAFP and uBMK with point mass has been fitted using maximum likelihood method. Numerical results show that the sAFP data and uBMK data are very well described by proposed model. A tree-structured sequential test can be optimized by selecting the cutoffs. Bootstrap simulations also show the robust classification results with the optimal cutoff.

Recent advances in genome-editing techniques have made it possible to modify any desired DNA sequence by employing programmable nucleases. These next-generation genome-modifying tools are the ideal candidates for therapeutic applications, especially for the treatment of genetic disorders like sickle cell disease (SCD). SCD is an inheritable monogenic disorder which is caused by a point mutation in the β-globin gene. Substantial success has been achieved in the development of supportive therapeutic strategies for SCD, but unfortunately there is still a lack of long-term universal cure. The only existing curative treatment is based on allogeneic stem cell transplantation from healthy donors; however, this treatment is applicable to a limited number of patients only. Hence, a universally applicable therapy is highly desirable. In this review, we will discuss the three programmable nucleases that are commonly used for genome-editing purposes: zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9). We will continue by exemplifying uses of these methods to correct the sickle cell mutation. Additionally, we will present induction of fetal globin expression as an alternative approach to cure sickle cell disease. We will conclude by comparing the three methods and explaining the concerns about their use in therapy.

Aim: Aberrant methylation of the promoter, P2, and the first exon, E1, regions of the tumor suppressor gene RASSF1A, have been associated with hepatocellular carcinoma (HCC), albeit with poor specificity. This study analyzed the methylation profiles of P1, P2 and E1 regions of the gene to identify the region of which methylation most specifically corresponds to HCC and to evaluate the potential of this methylated region as a biomarker in urine for HCC screening., Methods: Bisulfite DNA sequencing and quantitative methylation-specific polymerase chain reaction assays were performed to compare methylation of the 56 CpG sites in regions P1, P2 and E1 in DNA isolated from normal, hepatitic, cirrhotic, adjacent non-HCC, and HCC liver tissue and urine samples for the characterization of hypermethylation of the RASSF1A gene as a biomarker for HCC screening., Results: In tissue, comparing HCC (n = 120) with cirrhosis and hepatitis together (n = 70), methylation of P1 had an area under the receiver operating characteristics curve (AUROC) of 0.90, whereas methylation of E1 and P2 had AUROC of 0.84 and 0.72, respectively. At 90% sensitivity, specificity for P1 methylation was 72.9% versus 38.6% for E1 and 27.1% for P2. Methylated P1 DNA was detected in urine in association with cirrhosis and HCC. It had a sensitivity of 81.8% for α-fetoprotein negative HCC., Conclusion: Among the three regions analyzed, methylation of P1 is the most specific for HCC and holds great promise as a DNA marker in urine for screening of cirrhosis and HCC., (Published 2014. This article is a U.S. Government work and is in the public domain in the USA.)

Background: Neurosurgical resection and whole-brain radiation therapy (WBRT) are accepted treatments for single and oligometastatic cancer to the brain. To avoid the decline in neurocognitive function (NCF) linked to WBRT, the authors conducted a prospective, multicenter, phase 2 study to determine whether surgery and carmustine wafers (CW), while deferring WBRT, could preserve NCF and achieve local control (LC)., Methods: NCF and LC were measured in 59 patients who underwent resection and received CW for a single (83%) or dominant (oligometastatic, 2 to 3 lesions) metastasis and received stereotactic radiosurgery (SRS) for tiny nodules not treated with resection plus CW. Preservation of NCF was defined as an improvement or a decline ≤ 1 standard deviation from baseline in 3 domains: memory, executive function, and fine motor skills, evaluated at 2-month intervals., Results: Significant improvements in executive function and memory occurred throughout the 1-year follow-up. Preservation or improvement of NCF occurred in all 3 domains for the majority of patients at each of the 2-month intervals. NCF declined in only 1 patient. The chemowafers were well tolerated, and serious adverse events were reversible. There was local recurrence in 28% of the patients at 1-year follow-up., Conclusions: Patients with brain metastases had improvements in their cognitive trajectory, especially memory and executive function, after treatment with resection plus CW. The rate of LC (78%) was comparable to historic rates of surgery with WBRT and superior to reports of WBRT alone. For patients who undergo resection for symptomatic or large-volume metastasis or for tissue diagnosis, the addition of CW can be considered as an option., (© 2013 The Authors. Cancer published by Wiley Periodicals, Inc. on behalf of American Cancer Society.)

Spinal reconstruction of the L5 vertebrae after tumor resection remains a challenge. Complex resection followed by circumferential fixation in the same setting, or in staged fashion, is often employed. The added operative time associated with this method potentially increases morbidity and mortality in an inherently high-risk procedure and anatomy in the lumbosacral area makes reconstruction more challenging. The authors describe a technique involving L5 vertebrectomy, placement of an expandable cage, and anterolateral L4-S1 screw fixation via a one-stage, one-position, anterolateral retroperitoneal approach. Two illustrative cases are presented along with the authors overall experience in L5 tumor operations. We believe that this is a feasible reconstructive option after tumor resection in lower lumbar metastatic spine disease. The approach may be also utilized in combined anteroposterior (two-stage) procedures in primary malignant tumors or oligometastatic disease., (Copyright © 2011 Wiley-Liss, Inc.)

Vascular smooth muscle cell (VSMC) migration is an important cellular event in multiple vascular diseases, including atherosclerosis, restenosis, and transplant vasculopathy. Little is known regarding the effects of anti-inflammatory interleukins on VSMC migration. This study tested the hypothesis that an anti-inflammatory Th2 interleukin, interleukin-19 (IL-19), could decrease VSMC motility. IL-19 significantly decreased platelet-derived growth factor (PDGF)-stimulated VSMC chemotaxis in Boyden chambers and migration in scratch wound assays. IL-19 significantly decreased VSMC spreading in response to PDGF. To determine the molecular mechanism(s) for these cellular effects, we examined the effect of IL-19 on activation of proteins that regulate VSMC cytoskeletal dynamics and locomotion. IL-19 decreased PDGF-driven activation of several cytoskeletal regulatory proteins that play an important role in smooth muscle cell motility, including heat shock protein-27 (HSP27), myosin light chain (MLC), and cofilin. IL-19 decreased PDGF activation of the Rac1 and RhoA GTPases, important integrators of migratory signals. IL-19 was unable to inhibit VSMC migration nor was able to inhibit activation of cytoskeletal regulatory proteins in VSMC transduced with a constitutively active Rac1 mutant (RacV14), suggesting that IL-19 inhibits events proximal to Rac1 activation. Together, these data are the first to indicate that IL-19 can have important inhibitory effects on VSMC motility and activation of cytoskeletal regulatory proteins. This has important implications for the use of anti-inflammatory cytokines in the treatment of vascular occlusive disease.

Objective: The causes of failure after an initial Gamma procedure were studied, along with imaging and clinical outcomes, in a series of 140 patients with cerebral arteriovenous malformations (AVMs) treated with repeat Gamma Knife surgery (GKS)., Methods: Causes of initial treatment failure included inaccurate nidus definition in 14 patients, failure to fill part of the nidus as a result of hemodynamic factors in 16, recanalization of embolized AVM compartments in 6, and suboptimal dose (<20 Gy) in 23. Nineteen patients had repeat GKS for subtotal obliteration of AVMs. In 62 patients, the AVM failed to obliterate despite correct target definition and adequate dose. At the time of retreatment, the nidus volume ranged from 0.1 to 6.9 cm3 (mean, 1.4 cm3), and the mean prescription dose was 20.3 Gy., Results: Repeat GKS yielded a total angiographic obliteration in 77 patients (55%) and subtotal obliteration in 9 (6.4%). In 38 patients (27.1%), the AVMs remained patent, and in 16 patients (11.4%), no flow voids were observed on magnetic resonance imaging. Clinically, 126 patients improved or remained stable, and 14 experienced deterioration (8 resulting from a rebleed, 2 caused by persistent arteriovenous shunting, and 4 related to radiation-induced changes)., Conclusion: By using repeat GKS, we achieved a 55% angiographic cure rate. Although radiation-induced changes as visualized on magnetic resonance imaging occurred in 48 patients (39%), only 4 patients (3.6%) developed permanent neurological deficits. These findings may be useful in deciding the management of AVMs in whom total obliteration after initial GKS was not achieved.

Endothelial cell (EC) activation plays a key role in vascular inflammation, thrombosis, and angiogenesis. Allograft inflammatory factor-1 (AIF-1) is a cytoplasmic, calcium-binding, inflammation-responsive scaffold protein that has been implicated in the regulation of inflammation. The expression and function of AIF-1 in EC is uncharacterized, and the purpose of this study was to characterize AIF-1 expression and function in ECs. AIF-1 expression colocalized with CD31-positive ECs in neointima of inflamed human arteries but not normal arteries. AIF-1 is detected at low levels in unstimulated EC, but expression can be increased in response to serum and soluble factors. Stable transfection of AIF-1 small interfering RNA (siRNA) in ECs reduced AIF-1 protein expression by 73% and significantly reduced EC proliferation and migration (P < 0.05 and 0.001). Rescue of AIF-1 expression restored both proliferation and migration of siRNA-expressing ECs, and AIF-1 overexpression enhanced both of these activities, suggesting a strong association between AIF-1 expression and EC activation. Activation of mitogen-activated protein kinase p44/42 and PAK1 was significantly reduced in siRNA ECs challenged with inflammatory stimuli. Reduction of AIF-1 expression did not decrease EC tube-like structure or microvessel formation from aortic rings, but overexpression of AIF-1 did significantly increase the number and complexity of these structures. These data indicate that AIF-1 expression plays an important role in signal transduction and activation of ECs and may also participate in new vessel formation.

Possible modes of deactivation of Jacobsen's Co-salen catalyst during the hydrolytic kinetic resolution (HKR) of epichlorohydrin were explored by UV-vis spectroscopy, X-ray absorption spectroscopy, and electrospray ionization mass spectrometry, combined with recycling studies. Although an active Co(III)-salen catalyst deactivated substantially after multiple cycles without regeneration, the catalyst maintained its +3 oxidation state throughout the runs. Thus, deactivation of Co-salen during HKR was not the result of Co reduction. The mass spectrum of a deactivated material showed that catalyst dimerization does not account for the loss of activity. Results from various catalyst pretreatment tests, as well as from catalysts containing various counterions (acetate, tosylate, chloride, iodide) indicated that the rate of addition of the Co-salen counterions to epoxide forming Co-OH during the reaction correlated with deactivation. The extent of counterion addition to epoxide was influenced by the exposure time and the nucleophilicity of the counterion. An oligo(cyclooctene)-supported Co-OAc salen catalyst, which was 25 times more active than the standard Co-salen catalyst, was recycled multiple times with negligible deactivation.