DNA damage predicts prognosis and treatment response in colorectal liver metastases superior to immunogenic cell death and T cells

Preclinical models indicate that DNA damage induces type I interferon (IFN), which is crucial for the induction of an anti-tumor immune response. In human cancers, however, the association between DNA damage and an immunogenic cell death (ICD), including the release and sensing of danger signals, the subsequent ER stress response and a functional IFN system, is less clear. Methods: Neoadjuvant-treated colorectal liver metastases (CLM) patients, undergoing liver resection in with a curative intent, were retrospectively enrolled in this study (n=33). DNA damage (gammaH2AX), RNA and DNA sensors (RIG-I, DDX41, cGAS, STING), ER stress response (p-PKR, p-eIF2alpha, CALR), type I and type II IFN- induced proteins (MxA, GBP1), mature dendritic cells (CD208), and cytotoxic and memory T cells (CD3, CD8, CD45RO) were investigated by an immunohistochemistry whole-slide tissue scanning approach and further correlated with recurrence-free survival (RFS), overall survival (OS), radiographic and pathologic therapy response. Results: gammaH2AX is a negative prognostic marker for RFS (HR 1.32, 95% CI 1.04-1.69, p=0.023) and OS (HR 1.61, 95% CI 1.23-2.11, p<0.001). A model comprising of DDX41, STING and p-PKR predicts radiographic therapy response (AUC=0.785, p=0.002). gammaH2AX predicts prognosis superior to the prognostic value of CD8. CALR positively correlates with GBP1, CD8 and cGAS. A model consisting of gammaH2AX, p-eIF2alpha, DDX41, cGAS, CD208 and CD45RO predicts pathological therapy response (AUC=0.944, p<0.001). Conclusion: In contrast to preclinical models, DNA damage inversely correlated with ICD and its associated T cell infiltrate and potentially serves as a therapeutic target in CLM

Selective Denervation of the Facial Dermato-Muscular Complex in the Rat: Experimental Model and Anatomical Basis

The facial dermato-muscular system consists of highly specialized muscles tightly adhering to the overlaying skin and thus form a complex morphological conglomerate. This is the anatomical and functional basis for versatile facial expressions, which are essential for human social interaction. The neural innervation of the facial skin and muscles occurs via branches of the trigeminal and facial nerves. These are also the most commonly pathologically affected cranial nerves, often requiring surgical treatment. Hence, experimental models for researching these nerves and their pathologies are highly relevant to study pathophysiology and nerve regeneration. Experimental models for the distinctive investigation of the complex afferent and efferent interplay within facial structures are scarce. In this study, we established a robust surgical model for distinctive exploration of facial structures after complete elimination of afferent or efferent innervation in the rat. Animals were allocated into two groups according to the surgical procedure. In the first group, the facial nerve and in the second all distal cutaneous branches of the trigeminal nerve were transected unilaterally. All animals survived and no higher burden was caused by the procedures. Whisker pad movements were documented with video recordings 4 weeks after surgery and showed successful denervation. Whole-mount immunofluorescent staining of facial muscles was performed to visualize the innervation pattern of the neuromuscular junctions. Comprehensive quantitative analysis revealed large differences in afferent axon counts in the cutaneous branches of the trigeminal nerve. Axon number was the highest in the infraorbital nerve (28,625 ± 2,519), followed by the supraorbital nerve (2,131 ± 413), the mental nerve (3,062 ± 341), and the cutaneous branch of the mylohyoid nerve (343 ± 78). Overall, this surgical model is robust and reliable for distinctive surgical deafferentation or deefferentation of the face. It may be used for investigating cortical plasticity, the neurobiological mechanisms behind various clinically relevant conditions like facial paralysis or trigeminal neuralgia as well as local anesthesia in the face and oral cavity

Escherichia coli Frameshift Mutation Rate Depends on the Chromosomal Context but Not on the GATC Content Near the Mutation Site

Different studies have suggested that mutation rate varies at different positions in the genome. In this work we analyzed if the chromosomal context and/or the presence of GATC sites can affect the frameshift mutation rate in the Escherichia coli genome. We show that in a mismatch repair deficient background, a condition where the mutation rate reflects the fidelity of the DNA polymerization process, the frameshift mutation rate could vary up to four times among different chromosomal contexts. Furthermore, the mismatch repair efficiency could vary up to eight times when compared at different chromosomal locations, indicating that detection and/or repair of frameshift events also depends on the chromosomal context. Also, GATC sequences have been proved to be essential for the correct functioning of the E. coli mismatch repair system. Using bacteriophage heteroduplexes molecules it has been shown that GATC influence the mismatch repair efficiency in a distance- and number-dependent manner, being almost nonfunctional when GATC sequences are located at 1 kb or more from the mutation site. Interestingly, we found that in E. coli genomic DNA the mismatch repair system can efficiently function even if the nearest GATC sequence is located more than 2 kb away from the mutation site. The results presented in this work show that even though frameshift mutations can be efficiently generated and/or repaired anywhere in the genome, these processes can be modulated by the chromosomal context that surrounds the mutation site

Identification of methylated deoxyadenosines in vertebrates reveals diversity in DNA modifications.

Methylation of cytosine deoxynucleotides generates 5-methylcytosine (m(5)dC), a well-established epigenetic mark. However, in higher eukaryotes much less is known about modifications affecting other deoxynucleotides. Here, we report the detection of N(6)-methyldeoxyadenosine (m(6)dA) in vertebrate DNA, specifically in Xenopus laevis but also in other species including mouse and human. Our methylome analysis reveals that m(6)dA is widely distributed across the eukaryotic genome and is present in different cell types but is commonly depleted from gene exons. Thus, direct DNA modifications might be more widespread than previously thought.M.J.K. was supported by the Long-Term Human Frontiers Fellowship (LT000149/2010-L), the Medical Research Council grant (G1001690), and by the Isaac Newton Trust Fellowship (R G76588). The work was sponsored by the Biotechnology and Biological Sciences Research Council grant BB/M022994/1 (J.B.G. and M.J.K.). The Gurdon laboratory is funded by the grant 101050/Z/13/Z (J.B.G.) from the Wellcome Trust, and is supported by the Gurdon Institute core grants, namely by the Wellcome Trust Core Grant (092096/Z/10/Z) and by the Cancer Research UK Grant (C6946/A14492). C.R.B. and G.E.A. are funded by the Wellcome Trust Core Grant. We are grateful to D. Simpson and R. Jones-Green for preparing X. laevis eggs and oocytes, F. Miller for providing us with M. musculus tissue, T. Dyl for X. laevis eggs and D. rerio samples, and to Gurdon laboratory members for their critical comments. We thank U. Ruether for providing us with M. musculus kidney DNA (Entwicklungs- und Molekularbiologie der Tiere, Heinrich Heine Universitaet Duesseldorf, Germany). We also thank J. Ahringer, S. Jackson, A. Bannister and T. Kouzarides for critical input and advice, M. Sciacovelli and E. Gaude for suggestions.This is the author accepted manuscript. The final version is available from Nature Publishing Group via http://dx.doi.org/10.1038/nsmb.314

Deficient Dopamine D2 Receptor Function Causes Renal Inflammation Independently of High Blood Pressure

Renal dopamine receptors participate in the regulation of blood pressure. Genetic factors, including polymorphisms of the dopamine D2 receptor gene (DRD2) are associated with essential hypertension, but the mechanisms of their contribution are incompletely understood. Mice lacking Drd2 (D2−/−) have elevated blood pressure, increased renal expression of inflammatory factors, and renal injury. We tested the hypothesis that decreased dopamine D2 receptor (D2R) function increases vulnerability to renal inflammation independently of blood pressure, is an immediate cause of renal injury, and contributes to the subsequent development of hypertension. In D2−/− mice, treatment with apocynin normalized blood pressure and decreased oxidative stress, but did not affect the expression of inflammatory factors. In mouse RPTCs Drd2 silencing increased the expression of TNFα and MCP-1, while treatment with a D2R agonist abolished the angiotensin II-induced increase in TNF-α and MCP-1. In uni-nephrectomized wild-type mice, selective Drd2 silencing by subcapsular infusion of Drd2 siRNA into the remaining kidney produced the same increase in renal cytokines/chemokines that occurs after Drd2 deletion, increased the expression of markers of renal injury, and increased blood pressure. Moreover, in mice with two intact kidneys, short-term Drd2 silencing in one kidney, leaving the other kidney undisturbed, induced inflammatory factors and markers of renal injury in the treated kidney without increasing blood pressure. Our results demonstrate that the impact of decreased D2R function on renal inflammation is a primary effect, not necessarily associated with enhanced oxidant activity, or blood pressure; renal damage is the cause, not the result, of hypertension. Deficient renal D2R function may be of clinical relevance since common polymorphisms of the human DRD2 gene result in decreased D2R expression and function

Axonal mapping of the motor cranial nerves

Basic behaviors, such as swallowing, speech, and emotional expressions are the result of a highly coordinated interplay between multiple muscles of the head. Control mechanisms of such highly tuned movements remain poorly understood. Here, we investigated the neural components responsible for motor control of the facial, masticatory, and tongue muscles in humans using specific molecular markers (ChAT, MBP, NF, TH). Our findings showed that a higher number of motor axonal population is responsible for facial expressions and tongue movements, compared to muscles in the upper extremity. Sensory axons appear to be responsible for neural feedback from cutaneous mechanoreceptors to control the movement of facial muscles and the tongue. The newly discovered sympathetic axonal population in the facial nerve is hypothesized to be responsible for involuntary control of the muscle tone. These findings shed light on the pivotal role of high efferent input and rich somatosensory feedback in neuromuscular control of finely adjusted cranial systems

Global patient outcomes after elective surgery: prospective cohort study in 27 low-, middle- and high-income countries.

BACKGROUND: As global initiatives increase patient access to surgical treatments, there remains a need to understand the adverse effects of surgery and define appropriate levels of perioperative care. METHODS: We designed a prospective international 7-day cohort study of outcomes following elective adult inpatient surgery in 27 countries. The primary outcome was in-hospital complications. Secondary outcomes were death following a complication (failure to rescue) and death in hospital. Process measures were admission to critical care immediately after surgery or to treat a complication and duration of hospital stay. A single definition of critical care was used for all countries. RESULTS: A total of 474 hospitals in 19 high-, 7 middle- and 1 low-income country were included in the primary analysis. Data included 44 814 patients with a median hospital stay of 4 (range 2-7) days. A total of 7508 patients (16.8%) developed one or more postoperative complication and 207 died (0.5%). The overall mortality among patients who developed complications was 2.8%. Mortality following complications ranged from 2.4% for pulmonary embolism to 43.9% for cardiac arrest. A total of 4360 (9.7%) patients were admitted to a critical care unit as routine immediately after surgery, of whom 2198 (50.4%) developed a complication, with 105 (2.4%) deaths. A total of 1233 patients (16.4%) were admitted to a critical care unit to treat complications, with 119 (9.7%) deaths. Despite lower baseline risk, outcomes were similar in low- and middle-income compared with high-income countries. CONCLUSIONS: Poor patient outcomes are common after inpatient surgery. Global initiatives to increase access to surgical treatments should also address the need for safe perioperative care. STUDY REGISTRATION: ISRCTN5181700