Ocular surface information seen from the somatosensory thalamus and cortex.

Ocular Surface (OS) somatosensory innervation detects external stimuli producing perceptions, such as pain or dryness, the most relevant symptoms in many OS pathologies. Nevertheless, little is known about the central nervous system circuits involved in these perceptions, and how they integrate multimodal inputs in general. Here, we aim to describe the thalamic and cortical activity in response to OS stimulation of different modalities. Electrophysiological extracellular recordings in anaesthetized rats were used to record neural activity, while saline drops at different temperatures were applied to stimulate the OS. Neurons were recorded in the ophthalmic branch of the trigeminal ganglion (TG, 49 units), the thalamic VPM-POm nuclei representing the face (Th, 69 units) and the primary somatosensory cortex (S1, 101 units). The precise locations for Th and S1 neurons receiving OS information are reported here for the first time. Interestingly, all recorded nuclei encode modality both at the single neuron and population levels, with noxious stimulation producing a qualitatively different activity profile from other modalities. Moreover, neurons responding to new combinations of stimulus modalities not present in the peripheral TG subsequently appear in Th and S1, being organized in space through the formation of clusters. Besides, neurons that present higher multimodality display higher spontaneous activity. These results constitute the first anatomical and functional characterization of the thalamocortical representation of the OS. Furthermore, they provide insight into how information from different modalities gets integrated from the peripheral nervous system into the complex cortical networks of the brain. KEY POINTS: Anatomical location of thalamic and cortical ocular surface representation. Thalamic and cortical neuronal responses to multimodal stimulation of the ocular surface. Increasing functional complexity along trigeminal neuroaxis. Proposal of a new perspective on how peripheral activity shapes central nervous system function.

DNA damage, alterations in the expression of antioxidant enzyme genes and in the histoarchitecture of gill cells of zebrafish exposed to 17-α-ethinylestradiol.

Endocrine disruptors, such as estrogen, are chemical substances with the potential to alter the hormonal balance of organisms. Their origin can be natural or artificial, and they can act at very low doses. The estrogen 17α-ethinylestradiol (EE2) is used worldwide as an oral contraceptive and is a potential contaminant in aquatic ecosystems. It is well documented that these environmental pollutants can act directly or indirectly on the reproductive system, impairing development and fertility. However, little is known about the alteration of the cell oxidative status induced by EE2. The main objective of this study was to evaluate the effect on the gill cells of adult zebrafish exposed in vivo to EE2, analyzing cell histology, DNA damage and the expression levels of genes encoding the main enzymes involved in oxidative stress pathways. The histological study showed that EE2 produces moderate to high damage to the gill tissue, an increase in gill cell DNA damage and the mRNA levels of the genes corresponding to the manganese superoxide dismutase (Mn-sod) and catalase (cat) after exposure to 5 ng/L EE2. The results indicate that EE2 causes tissue alterations, DNA damage and oxidative stress. EE2 produced important alterations in the gills, a fundamental organ for the survival of fish. There is a clear need for further research on the ecological consequences of EDCs on non-target organisms.

Green synthesis of gold nanoparticles via Moringa oleifera seed extract: antioxidant, antibacterial and anticarcinogenic activity on lung cancer.

Plant-mediated biosynthesis of nanoparticles is a green method that allows synthesis in one-pot process. Synthesis of gold nanoparticles with plant extracts has gained interest in the field of biomedicine due to its variety of applications. This study presents the synthesis via green chemistry of gold nanoparticles (AuNPs) using the methanol extract of Moringa oleifera seeds. The AuNPs were synthesized at room temperature. UV-Vis spectroscopy confirmed the formation of AuNPs by identifying the surface plasmon resonance located at 546 nm. TEM analysis shows spherical nanoparticles. FTIR analysis demonstrated the presence of specific bioactive molecules responsible for the Au3+ ion reduction process. The antioxidant activity of the nanoparticles was evaluated on the stabilization of the DPPH radical (1,1-diphenyl-2-picrylhydrazyl, 2,2-diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl). The antimicrobial activity analysis was developed by broth microdilution method at different concentrations against Escherichia coli and Staphylococcus aureus. Minimum inhibitory concentration were 400 µg/mL and 200 µg/mL, respectively. A549 lung cancer cell proliferation was measured according to the MTT protocol, indicating a dose-dependent response and a IC50 of 163.9 ± 13.27 µg/mL. The AuNPs synthesized using M. oleifera seeds showed promise as active materials for antimicrobial or anticancer products.

Biodegradable nanofiber-based piezoelectric transducer.

Piezoelectric materials, a type of "smart" material that generates electricity while deforming and vice versa, have been used extensively for many important implantable medical devices such as sensors, transducers, and actuators. However, commonly utilized piezoelectric materials are either toxic or nondegradable. Thus, implanted devices employing these materials raise a significant concern in terms of safety issues and often require an invasive removal surgery, which can damage directly interfaced tissues/organs. Here, we present a strategy for materials processing, device assembly, and electronic integration to 1) create biodegradable and biocompatible piezoelectric PLLA [poly(l-lactic acid)] nanofibers with a highly controllable, efficient, and stable piezoelectric performance, and 2) demonstrate device applications of this nanomaterial, including a highly sensitive biodegradable pressure sensor for monitoring vital physiological pressures and a biodegradable ultrasonic transducer for blood-brain barrier opening that can be used to facilitate the delivery of drugs into the brain. These significant applications, which have not been achieved so far by conventional piezoelectric materials and bulk piezoelectric PLLA, demonstrate the PLLA nanofibers as a powerful material platform that offers a profound impact on various medical fields including drug delivery, tissue engineering, and implanted medical devices.

The Contribution of TRPA1 to Corneal Thermosensitivity and Blink Regulation in Young and Aged Mice.

The role of TRPA1 in the thermosensitivity of the corneal cold thermoreceptor nerve endings was studied in young and aged mice. The contribution of the TRPA1-dependent activity to basal tearing and thermally-evoked blink was also explored. The corneal cold thermoreceptors' activity was recorded extracellularly in young (5-month-old) and aged (18-month-old) C57BL/6WT (WT) and TRPA1-/- knockout (TRPA1-KO) mice at basal temperature (34 °C) and during cooling (15 °C) and heating (45 °C) ramps. The blink response to cold and heat stimulation of the ocular surface and the basal tearing rate were also measured in young animals using orbicularis oculi muscle electromyography (OOemg) and phenol red threads, respectively. The background activity at 34 °C and the cooling- and heating-evoked responses of the cold thermoreceptors were similar in WT and TRPA1-KO animals, no matter the age. Similar to the aged WT mice, in the young and aged TRPA1-KO mice, most of the cold thermoreceptors presented low frequency background activity, a low cooling threshold, and a sluggish response to heating. The amplitude and duration of the OOemg signals correlated with the magnitude of the induced thermal change in the WT but not in the TRPA1-KO mice. The basal tearing was similar in the TRPA1-KO and WT mice. The electrophysiological data suggest that the TRPA1-dependent nerve activity, which declines with age, contributes to detecting the warming of the ocular surface and also to integrating the thermally-evoked reflex blink.

Cyclosporine A Decreases Dryness-Induced Hyperexcitability of Corneal Cold-Sensitive Nerve Terminals.

Cyclosporine A (CsA) is used for the treatment of dry eye (DE) with good clinical results, improving tear secretion and decreasing subjective symptoms. These effects are attributed to the improved tear film dynamics, but there are no data on the effect of CsA on the abnormal sensory nerve activity characteristic in DE. Our purpose was to evaluate the CsA effect on the enhanced activity of corneal cold thermoreceptors in a tear-deficient DE animal model using in vitro extracellular recording of cold thermoreceptors nerve terminal impulses (NTIs) before and in the presence of CsA. NTI shape was also analyzed. Blinking frequency and tearing rate were also measured in awake animals before and after topical CsA. CsA increased the tearing and blinking of treated animals. CsA significantly decreased the peak response to cold of cold thermoreceptors. Neither their spontaneous NTIs discharge rate nor their cooling threshold were modified. CsA also seemed to reverse some of the changes in NTI shape induced by tear deficiency. These data suggest that, at least in part, the beneficial clinical effects of CsA in DE can be attributed to a direct effect on sensory nerve endings, although the precise mechanisms underlying this effect need further studies to be fully clarified.

Clinical features and genetic biomarkers associated with different phenotypes of systemic lupus erythematosus in Paraguayan patients.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by a heterogeneous clinical picture that makes the diagnosis and follow-up of these patients difficult. This study aimed to identify correlations between clinical, immunological, and genetic biomarkers and clinical manifestations in SLE. A retrospective study of data from medical records and immunological and genetic studies of SLE patients in Paraguay was carried out. A descriptive analysis was performed based on the type of variable. Human leukocyte antigen (HLA) allele frequencies (DPA1, DPB1, DQA1, DQB1, and DRB1) were calculated, and univariate logistic regression analyses were performed between each of the explanatory variables and the presence or absence of each phenotype. Odds ratios, 95% confidence intervals, and p values were recorded. Associations with p<0.05 were considered statistically significant. 104 SLE patients were included: 86% were female, with a mean age of 32.80±10.36 years. An association was identified between anti-double stranded DNA (anti-dsDNA) and the presence of the renal phenotype and between anti-dsDNA and the absence of the joint and hematological phenotypes. Immunoglobulin M isotype rheumatoid factor was associated with the absence of a renal phenotype. HLA-DQB1*02:02 and HLA-DRB1*07:01 were associated with the cutaneous phenotype. An association was identified between age at disease onset over 30 years and the presence of the joint phenotype. No other associations were identified. Potential clinical, immunological, and genetic biomarkers of phenotypes have been identified in SLE Paraguayan patients.

Self-assembled materials from cellulose nanocrystals conjugated with a thermotropic liquid crystalline moiety.

Manipulating molecular and supramolecular interactions within cellulose nanocrystals (CNCs) to introduce different levels of assemblies combined with multiple functionalities is required for the development of degradable smart materials from renewable resources. To attain hierarchical structures and stimuli-responsive properties, a new class of liquid crystalline cellulosic hybrid materials is synthesized. Herein, main-chain rigid-rod-like oxidized cellulose (CNC-COOH) is prepared from a Cellulose Whatman filter paper (Cellulose W.P.) by acid hydrolysis and oxidized using 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO). Thermotropic LC molecule, 4-cyano-4'-hydroxybiphenyl with a 12-methylene spacer (CB12-OH) is grafted onto the carboxylic acid group of CNC-COOH via Steglich esterification. The liquid crystalline functionalized CNCs cellulose nanocrystals (CNC-COO-CB12) are readily soluble in DMSO and ionic liquids. The extent of functionalization and structure of CNC-COO-CB12 are confirmed by solution-state 1H NMR and supported by other characterization techniques. We investigate the interplay of liquid crystalline orientational order of CNCs and cyanobiphenyl (CB12), and the supramolecular hydrogen bonding of CNCs within CNC-COO-CB12 and compare it with CNC-COOH. The introduction of thermotropic CB12 side chains onto rigid-rod CNCs shows the exclusive formation of smectic mesophases from the assemblies of CB12 with the absence of the cholesteric mesophase typically observed from CNC-COOH as verified by temperature-controlled SAXS (T-SAXS). This is further verified by UV-visible and SEM studies that show CNC-COO-CB12 forms smectic domains while CNC-COOH forms a visible light reflecting cholesteric mesophase in dried films. Thus, the interplay of liquid crystalline order of CNCs and CB12 and supramolecular hydrogen bonding of CNCs results in ordered, smectic-mesostructured CNCs for use in stimuli-responsive functional materials.

An Experimental Model of Neuro-Immune Interactions in the Eye: Corneal Sensory Nerves and Resident Dendritic Cells.

The cornea is an avascular connective tissue that is crucial, not only as the primary barrier of the eye but also as a proper transparent refractive structure. Corneal transparency is necessary for vision and is the result of several factors, including its highly organized structure, the physiology of its few cellular components, the lack of myelinated nerves (although it is extremely innervated), the tightly controlled hydration state, and the absence of blood and lymphatic vessels in healthy conditions, among others. The avascular, immune-privileged tissue of the cornea is an ideal model to study the interactions between its well-characterized and dense sensory nerves (easily accessible for both focal electrophysiological recording and morphological studies) and the low number of resident immune cell types, distinguished from those cells migrating from blood vessels. This paper presents an overview of the corneal structure and innervation, the resident dendritic cell (DC) subpopulations present in the cornea, their distribution in relation to corneal nerves, and their role in ocular inflammatory diseases. A mouse model in which sensory axons are constitutively labeled with tdTomato and DCs with green fluorescent protein (GFP) allows further analysis of the neuro-immune crosstalk under inflammatory and steady-state conditions of the eye.

A Study of Zn-Ca Nanocomposites and Their Antibacterial Properties.

This study aimed to develop Ca2+ doped ZnO nanoparticles (NPs) and investigate their antibacterial properties against microorganisms of dental interest. Zn-Ca NPs were synthesized by the sol-gel method with different concentrations of Ca2+ (1, 3, and 5 wt. %) and subsequently characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD), UV-vis spectroscopy and Fourier transform infrared spectroscopy (FT-IR). The Kirby-Bauer method was used to measure antibacterial effects. NPs showed the wurzite phase of ZnO and bandgap energies (Eg) from 2.99 to 3.04 eV. SEM analysis showed an average particle size of 80 to 160 nm. The treatments that presented the best antibacterial activity were Zn-Ca 3% and Zn-Ca 5%. ZnO NPs represent an alternative to generate and improve materials with antibacterial capacity for dental applications.

Origin of the Co-Seismic Variations of Elastic Properties in the Crust: Insight From the Laboratory.

Seismological observations highlighted that earthquakes are often followed by changes in elastic properties around the fault zone. Here, we studied the origin of these variations using stick-slip experiments on saw-cut granite samples presenting different degrees of bulk damage (i.e., microcracks). Stick-slip events were induced under triaxial compression configuration with continuous active ultrasonic measurements at confining pressures representative of upper crustal conditions (15-120 MPa). Both the P-wave velocity ( V P ) and amplitude ( A P ) showed drops, concurrently with stress drops, and had a non-monotonic dependence toward the fault's stress state. Our experimental results suggest that co-seismic changes in V P were mostly controlled by the elastic re-opening of microcracks in the bulk, rather than by co-seismic damage or the formation of fault gouge. Co-seismic changes in A P were controlled by a combination of elastic re-opening of microcracks in the bulk and inelastic processes (i.e., co-seismic damage and gouge formation and dilation).

The role of brain biopsy in the clinical management of HIV-related focal brain lesions.

OBJECTIVES: Up to 20% of HIV-related focal brain lesion (FBL) diagnoses cannot be determined without invasive procedures. In such cases, brain biopsy is an important step in the evaluation algorithm. The aims of this study were to describe the clinical outcomes of patients with FBL, the proportion of diagnoses confirmed by brain biopsies and their aetiologies, and to analyse the proportion of patients in whom the biopsy motivated a change in therapeutic management. METHODS: A retrospective cohort study was performed. The data from clinical records of patients with HIV-related FBL admitted between January 2005 and December 2015 were reviewed. RESULTS: A total of 137 patients were included in the study. The median age was 39 years [interquartile range (IQR) 33-44.5 years]. The median CD4 count was 54 cells/µL (IQR 21-124 cells/µL). Cerebral brain biopsy was performed in 21.16% of patients (29 of 137); 68.9% of these individuals (20 of 29) were diagnosed by histology, with results of central nervous system (CNS) lymphoma in 20.6% (six of 29), progressive multifocal leucoencephalopathy in 6.8% (two of 29), toxoplasmosis in 6.8% (two of 29), tuberculoma in 6.8% (two of 29), and other diagnoses in 27.6% (eight of 29). In nine patients, the histology was nonspecific. In 75.8% of patients (22 of 29), the result of the biopsy led to a change in the therapeutic management. We did not observe higher rates of mortality related to the procedure. Overall mortality at 30 and 90 days was similar in patients who were and were not biopsied. CONCLUSIONS: In this retrospective cohort study, cerebral biopsy was associated with significant adjustments in therapeutic management for a high percentage of patients.

Evolution After Anti-TNF Discontinuation in Patients With Inflammatory Bowel Disease: A Multicenter Long-Term Follow-Up Study.

OBJECTIVES: The aims of this study were to assess the risk of relapse after discontinuation of anti-tumor necrosis factor (anti-TNF) drugs in patients with inflammatory bowel disease (IBD), to identify the factors associated with relapse, and to evaluate the overcome after retreatment with the same anti-TNF in those who relapsed. METHODS: This was a retrospective, observational, multicenter study. IBD patients who had been treated with anti-TNFs and in whom these drugs were discontinued after clinical remission was achieved were included. RESULTS: A total of 1,055 patients were included. The incidence rate of relapse was 19% and 17% per patient-year in Crohn's disease and ulcerative colitis patients, respectively. In both Crohn's disease and ulcerative colitis patients in deep remission, the incidence rate of relapse was 19% per patient-year. The treatment with adalimumab vs. infliximab (hazard ratio (HR)=1.29; 95% confidence interval (CI)=1.01-1.66), elective discontinuation of anti-TNFs (HR=1.90; 95% CI=1.07-3.37) or discontinuation because of adverse events (HR=2.33; 95% CI=1.27-2.02) vs. a top-down strategy, colonic localization (HR=1.51; 95% CI=1.13-2.02) vs. ileal, and stricturing behavior (HR=1.5; 95% CI=1.09-2.05) vs. inflammatory were associated with a higher risk of relapse in Crohn's disease patients, whereas treatment with immunomodulators after discontinuation (HR=0.67; 95% CI=0.51-0.87) and age (HR=0.98; 95% CI=0.97-0.99) were protective factors. None of the factors were predictive in ulcerative colitis patients. Retreatment of relapse with the same anti-TNF was effective (80% responded) and safe. CONCLUSIONS: The incidence rate of inflammatory bowel disease relapse after anti-TNF discontinuation is relevant. Some predictive factors of relapse after anti-TNF withdrawal have been identified. Retreatment with the same anti-TNF drug was effective and safe.

Linkage and association analysis of ADHD endophenotypes in extended and multigenerational pedigrees from a genetic isolate.

Attention-deficit/hyperactivity disorder (ADHD) is a heritable, chronic, neurodevelopmental disorder with serious long-term repercussions. Despite being one of the most common cognitive disorders, the clinical diagnosis of ADHD is based on subjective assessments of perceived behaviors. Endophenotypes (neurobiological markers that cosegregate and are associated with an illness) are thought to provide a more powerful and objective framework for revealing the underlying neurobiology than syndromic psychiatric classification. Here, we present the results of applying genetic linkage and association analyses to neuropsychological endophenotypes using microsatellite and single nucleotide polymorphisms. We found several new genetic regions linked and/or associated with these endophenotypes, and others previously associated to ADHD, for example, loci harbored in the LPHN3, FGF1, POLR2A, CHRNA4 and ANKFY1 genes. These findings, when compared with those linked and/or associated to ADHD, suggest that these endophenotypes lie on shared pathways. The genetic information provided by this study offers a novel and complementary method of assessing the genetic causes underpinning the susceptibility to behavioral conditions and may offer new insights on the neurobiology of the disorder.

Genetic diversity patterns of arbuscular mycorrhizal fungi associated with the mycoheterotroph Arachnitis uniflora Phil. (Corsiaceae).

Background and Aims: Arachnitis uniflora is a mycoheterotrophic plant that exploits arbuscular mycorrhizal fungi of neighbouring plants. We tested A. uniflora 's specificity towards fungi across its large latitudinal range, as well as the role of historical events and current environmental, geographical and altitudinal variables on fungal genetic diversity. Methods: Arachnitis uniflora mycorrhizas were sampled at 25 sites. Fungal phylogenetic relationships were reconstructed, genetic diversity was calculated and the main divergent lineages were dated. Phylogeographical analysis was performed with the main fungal clade. Fungal diversity correlations with environmental factors were investigated. Key Results: Glomeraceae fungi dominated, with a main clade that likely originated in the Upper Cretaceous and diversified in the Miocene. Two other arbuscular mycorrhizal fungal families not previously known to be targeted by A. uniflora were detected rarely and appear to be facultative associations. High genetic diversity, found in Bolivia and both northern and southern Patagonia, was correlated with temperature, rainfall and soil features. Conclusions: Fungal genetic diversity and its distribution can be explained by the ancient evolutionary history of the target fungi and by micro-scale environmental conditions with a geographical mosaic pattern.

Nanowire networks and hollow nanospheres of Ag-Au bimetallic alloys at room temperature.

Due to their physicochemical properties, metallic nanoalloys have potential applications in biomedicine, electrocatalysis and electrochemical sensors, among many other fields. New alternative procedures have emerged in order to reduce production costs and the use of toxic substances. In this study we present a novel low-toxicity synthesis method for the fabrication of nanowire networks (NWNs) and Ag-Au hollow nanospheres. The synthesis process is performed at room temperature without any sophisticated equipment, such as special cameras or furnaces, etc. Transmission electron microscopy showed that the NWNs contain random alloys with a diameter of between 10-13 nm. The radius for the hollow nanospheres is approximately located between 70-130 nm. The absorption bands in the UV-vis spectrum associated with the surface plasmon in Ag-Au bimetallic nanoparticles are highlighted at 385 nm for the NWNs and 643 nm for the hollow nanospheres. The study was performed with low-toxicity substances, such as rongalite, ascorbic acid and sucrose, and showed high efficiency for the fabrication of these types of nanostructures, as well as good stability for long periods of time.

Evaluation of efficacy of impregnated curtains in experimental hen houses as a phlebotomine control tool in northeast Argentina.

The aim of this study was to evaluate the effectiveness of insecticide-impregnated curtains against the entry of phlebotomine (Diptera: Psychodidae) flies into experimental slatted hen houses in an area endemic for American cutaneous leishmaniasis (ACL). Three treatments in experimental dwellings containing three chickens each were applied using, respectively, an impregnated curtain (IC), a non-impregnated curtain (NIC) and no curtain (NC). A control site without chickens (WC) was included. The study used permethrin at 0.05 g/m2 . During each month for 1 year, each experimental hen house randomly received all treatments. Phlebotomine sandflies were captured using REDILA BL traps placed inside the hen house. Significant differences in abundances of phlebotomine flies/trap/night were observed between treatments (χ2 = 17853.58, d.f. = 3, P < 0.0001): 59.7% of phlebotomines were captured in the NC treatment, 26.3% in the NIC treatment, 8.0% in the IC treatment and 6.1% in the WC condition. Nyssomyia whitmani (Antunes & Coutinho) was the most abundant species in all collections (89.9%). These results showed a lower abundance of phlebotomines in the experimental hen house in the IC condition than in the hen house in the NC condition (P < 0.05) and that the presence of NIC represents an effective physical barrier against phlebotomines (P < 0.05). Therefore, the use of curtains may be an alternative eco-friendly method for the prevention of indoor ACL transmission in slatted dwellings, which represent a common house type in northeast Argentina.

Using the rd1 mouse to understand functional and anatomical retinal remodelling and treatment implications in retinitis pigmentosa: A review.

Retinitis Pigmentosa (RP) reflects a range of inherited retinal disorders which involve photoreceptor degeneration and retinal pigmented epithelium dysfunction. Despite the multitude of genetic mutations being associated with the RP phenotype, the clinical and functional manifestations of the disease remain the same: nyctalopia, visual field constriction (tunnel vision), photopsias and pigment proliferation. In this review, we describe the typical clinical phenotype of human RP and review the anatomical and functional remodelling which occurs in RP determined from studies in the rd/rd (rd1) mouse. We also review studies that report a slowing down or show an acceleration of retinal degeneration and finally we provide insights on the impact retinal remodelling may have in vision restoration strategies.

Peritoneal Lavage in Complicated Acute Diverticulitis: Back to the Future.

PURPOSE: The aim of this study was to assess laparoscopic lavage in patients with acute diverticulitis. BACKGROUND: In recent times, laparoscopic peritoneal lavage has been considered a therapeutic alternative to standard resection procedures. In Hinchey III diverticulitis there is the possibility of avoiding ostomies or resection procedures. STUDY: This retrospective study was conducted in a single-care institution. RESULTS: Patients requiring emergency surgery for perforated diverticulitis and generalized peritonitis between March 2011 and May 2014 were identified from a prospective database. Seventy-two patients underwent surgery for diverticulitis. Forty-three patients presented with generalized peritonitis (Hinchey III) and 29 with gross fecal contamination (Hinchey IV). From the Hinchey III group, 17 patients (39.5%), with a median age of 56.8 years, were selected to undergo peritoneal lavage.Postoperative length of stay was 4.2 days, the rate of minor complications was 35%, that of major complications was 0%, and mortality rate was 0%. Over a median follow-up of 24.6 months none required reoperation. CONCLUSIONS: The number of patients selected for laparoscopic lavage in the management of perforated diverticulitis is increasing as it provides an alternative to Hartmann's procedure in emergency cases. This approach should be considered suitable for patients without important comorbidities, and only in centers experienced in laparoscopic surgery. It reduces the length of hospital stay. Morbidity and mortality are lower than in those in whom resection was considered necessary. Longer follow-up and results of other trials will be necessary to draw an adequate conclusion.

Detailed characterization of MLH1 p.D41H and p.N710D variants coexisting in a Lynch syndrome family with conserved MLH1 expression tumors.

Lynch syndrome (LS) is an autosomal dominant cancer-susceptibility disease caused by inactivating germline mutations in mismatch repair (MMR) genes. Variants of unknown significance (VUS) are often detected in mutational analysis of MMR genes. Here we describe a large family fulfilling Amsterdam I criteria carrying two rare VUS in the MLH1 gene: c.121G > C (p.D41H) and c.2128A > G (p.N710D). Collection of clinico-pathological data, multifactorial analysis, in silico predictions, and functional analyses were used to elucidate the clinical significance of the identified MLH1 VUS. Only the c.121G > C variant cosegregated with LS-associated tumors in the family. Diagnosed colorectal tumors were microsatellite unstable although immunohistochemical staining revealed no loss of MMR proteins expression. Multifactorial likelihood analysis classified c.2128A > G as a non-pathogenic variant and c.121G > C as pathogenic. In vitro functional tests revealed impaired MMR activity and diminished expression of c.121G > C. Accordingly, the N710 residue is located in the unconserved MLH1 C-terminal domain, whereas D41 is highly conserved and located in the ATPase domain. The obtained results will enable adequate genetic counseling of c.121G > C and c.2128A > G variant carriers and their families. Furthermore, they exemplify how cumulative data and comprehensive analyses are mandatory to refine the classification of MMR variants.