Association of CTLA-4 (AT)n Variants in Basal Cell Carcinoma and Squamous Cell Carcinoma Patients from Western Mexico.

The incidence of basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) is constantly increasing, becoming a significant health problem. CTLA-4 is a critical immune checkpoint, and it has been suggested that a variant of variable-number tandem repeat in the 3'-UTR of its gene, known as (AT)n, may be associated with a higher susceptibility to some cancers; however, little is known about genetic variants of the CTLA-4 gene in NMSC. To establish the association of this genetic variant in the CTLA-4 gene with the susceptibility of NMSC carcinogenesis in the Western Mexican population, samples from 150 BCC patients, 150 SCC patients, and 150 healthy individuals as the reference group (RG) were analyzed by endpoint PCR, followed by electrophoresis to genotype the samples. We found that the short-repeat 104/104 bp genotype may be a risk factor for BBC carcinogens (OR = 2.92, p = 0.03), whereas the long-repeat 106/106 bp genotype may be a protective factor for both BCC (OR = 0.13, p = 0.01) and SCC (OR = 0.32, p = 0.01) susceptibility. Our results show that in the Western Mexican population, long-repeat (AT)n variants in the CTLA-4 gene are associated with a protective factor in BCC and SCC. In contrast, short repeats are associated with a risk factor.

Uncovering the Expression Pattern of the Costimulatory Receptors ICOS, 4-1BB, and OX-40 in Exhausted Peripheral and Tumor-Infiltrating Natural Killer Cells from Patients with Cervical Cancer.

Cervical cancer (CC) poses a significant health burden, particularly in low- and middle-income countries. NK cells play a crucial role against CC; however, they can become exhausted and lose their cytotoxic capacity. This work explores the expression of costimulatory receptors (ICOS, 4-1BB, OX-40) in exhausted NK cells from CC patients. Peripheral blood and tumor biopsies were collected, and flow cytometry was used to evaluate the expression of costimulatory receptors in exhausted NK cells. There is an increase of peripheral exhausted NK cells (PD-1+TIGIT+) in CC patients; this subpopulation has a selectively increased expression of the costimulatory receptors ICOS and 4-1BB. An exhausted population is also highly increased in tumor-infiltrating NK cells, and it shows a dramatically increased expression of the costimulatory receptors ICOS (>15×) and 4-1BB (>10×) compared to peripheral NK cells. The exhausted cells, both in the periphery and in the tumor infiltrating lymphocytes (TILs), are also more likely than non-exhausted NK cell populations (PD-1-TIGIT-) to express these costimulatory receptors; increases ranging from 2.0× ICOS, 2.4× 4-1BB, and 2.6× OX-40 in CD56dim PBMCs to 1.5× ICOS, 5× 4-1BB, and 10× OX-40 in TILs were found. Our study demonstrates for the first time the increased expression of the costimulatory receptors ICOS, 4-1BB, and OX-40 in peripheral CD56dim, CD56bright, and tumor-infiltrating NK cells in CC. Targeting these receptors for stimulation could reverse exhaustion and be a promising immunotherapy strategy.

Feasibility and reliability of the Spanish version of the Youth Activity Profile questionnaire (YAP-Spain) in children and adolescents.

Considerable public health efforts across the globe have focused on promoting physical activity (PA) and minimizing sedentary behaviour (SB) in youths. However, it is important to have valid, reliable and feasible methods to assess these behaviours in youths. The purpose of this study was to analyse the feasibility and reliability of the Spanish version of the previously validated Youth Activity Profile questionnaire (YAP) in children and adolescents. The YAP-S is a 15-item self-report instrument designed to capture PA and SB in youths. A total of 604 children (5-12 years old) and 346 adolescents (12-17 years old) filled out the questionnaire twice (14 days apart). Feasibility was evaluated through required time and number of misunderstood questions by participants. The test-retest reliability was examined using the weighted kappa coefficient (κ) and intraclass correlation coefficient. The average time to complete the questionnaire was 28.85 ± 14.28 and 12.24 ± 9.84 minutes in children and adolescents, respectively. No misunderstanding of questions was reported. The questionnaire showed an adequate reliability for activity at school, out-of-school and sedentary behaviours (k = 0.61-0.77; ICC = 0.77-0.89) in children and adolescents. The YAP-S might be considered a feasible and reliable questionnaire for assessing PA and SB in Spanish children and adolescents.

Design of a Low-Cost Air Levitation System for Teaching Control Engineering.

Air levitation is the process by which an object is lifted without mechanical support in a stable position, by providing an upward force that counteracts the gravitational force exerted on the object. This work presents a low-cost lab implementation of an air levitation system, based on open solutions. The rapid dynamics makes it especially suitable for a control remote lab. Due to the system's nature, the design can be optimized and, with some precision trade-off, kept affordable both in cost and construction effort. It was designed to be easily adopted to be used as both a remote lab and as a hands-on lab.

Individualized exercise prescription in non-alcoholic steatohepatitis: a case report.

The effect of an individualized exercise programme on a non-alcoholic steatohepatitis case is presented. Before entering the programme the patient was treated with conventional recommendations on diet plus aerobic exercise during fourteen years, without major improvements of his analytical parameters.Two years after including him in a tailored exercise programme, aimed to fulfil the recommendations of the American College of Sports Medicine, his blood markers of liver dysfunction and cardiometabolic risk tended to improve. Consequently, our data support the idea that in non-alcoholic steatohepatitis the exercise-based therapeutic interventions should be individualized taking into account the cardio-respiratory and muscular fitness of the patient, rather than using generic behavioural recommendations.

Increase of IFN-γ and TNF-α production in CD107a + NK-92 cells co-cultured with cervical cancer cell lines pre-treated with the HO-1 inhibitor.

BACKGROUND: Natural killer (NK) cells eliminate virus-infected and tumor cells through the release of perforins and granzymes; they also produce Interferon gamma (IFN-γ) and Tumor necrosis factor alpha (TNF-α), which induce apoptosis in target cells. Many tumors express Heme oxygenase 1 (HO-1), and this expression has been associated with avoiding immunosuppression and apoptosis. In this work, HO-1+ Cervical cancer cell (CCC) lines were pre-treated with HO-1 inhibitor and we assessed whether this inhibition enhanced the sensitivity of CCC to NK cell activity. METHODS: We assessed the expression of HO-1 in HeLa, SiHa, and C-33A CCC by Flow cytometry (FC). CCC were pre-treated with SnPP or ZnPP HO-1 inhibitors. After that, NK-92 cells were co-cultured with HeLa, SiHa, and C-33A CCC pre-treated or not with HO-1 inhibitors, and the expression of IFN-γ, TNF-α, CD107a, Granzyme B, NKp44, NKp46, NKp30, and NKG2D was evaluated by FC. RESULTS: CCC lines HeLa, SiHa, and C-33A expressed HO-1. Inhibition of HO-1 in these cells increased the expression of IFN-γ and TNF-α in CD107a + NK-92 cells. We observed a reduction in the expression of NKG2D, NKp46, and NKp30 in NK cells co-cultured with HeLa and SiHa cells, and when HeLa and SiHa cells were pre-treated with the HO-1 inhibitors, the expression of NKG2D and NKp30 in NK cells was restored. We observed a similar effect in NK cells co-cultured with C-33A cells in NKp30 expression. CONCLUSION: Inhibition of HO-1 in CCC induces an increase in IFN-γ and TNF-α production in CD107a + NK-92 cells and restores NKG2D, NKp46 and NKp30 downmodulation in NK cells.

Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss.

BACKGROUND: The resistance of cancerous cells to chemotherapy remains the main limitation for cancer treatment at present. Doxorubicin (DOX) is a potent antitumor drug that activates the ubiquitin-proteasome system, but unfortunately it also activates the Nuclear factor kappa B (NF-кB) pathway leading to the promotion of tumor cell survival. MG132 is a drug that inhibits I kappa B degradation by the proteasome-avoiding activation of NF-кB. In this work, we studied the sensitizing effect of the MG132 proteasome inhibitor on the antitumor activity of DOX. METHODS: U937 human leukemia cells were treated with MG132, DOX, or both drugs. We evaluated proliferation, viability, apoptosis, caspase-3, -8, and -9 activity and cleavage, cytochrome c release, mitochondrial membrane potential, the Bcl-2 and Bcl-XL antiapoptotic proteins, senescence, p65 phosphorylation, and pro- and antiapoptotic genes. RESULTS: The greatest apoptosis percentage in U937 cells was obtained with a combination of MG132 + DOX. Likewise, employing both drugs, we observed a decrease in tumor cell proliferation and important caspase-3 activation, as well as mitochondrial membrane potential loss. Therefore, MG132 decreases senescence, p65 phosphorylation, and the DOX-induced Bcl-2 antiapoptotic protein. The MG132 + DOX treatment induced upregulation of proapoptotic genes BAX, DIABLO, NOXA, DR4, and FAS. It also induced downregulation of the antiapoptotic genes BCL-XL and SURVIVIN. CONCLUSION: MG132 sensitizes U937 leukemia cells to DOX-induced apoptosis, increasing its anti-leukemic effectiveness.

New Advances in the Study of CMTM6, a Focus on Its Novel Non-Canonical Cellular Locations, and Functions beyond Its Role as a PD-L1 Stabilizer.

CMTM6 is a membrane protein that acts as a regulator of PD-L1, maintaining its expression on the cell surface, and can prevent its lysosome-mediated degradation. It is unknown if CMTM6 is present in the plasma of patients with cervical cancer, and if it has non-canonical subcellular localizations in cell lines derived from cervical cancer. Our objective was to determine whether CMTM6 is found in plasma derived from cervical cancer patients and its subcellular localization in cell lines. Patient plasma was separated into exosome-enriched, exosome-free, and total plasma fractions. The levels of CMTM6 in each fraction were determined using ELISA and Western blot. Finally, for the cellular model, HeLa, SiHa, CaSki, and HaCaT were used; the subcellular locations of CMTM6 were determined using immunofluorescence and flow cytometry. Soluble CMTM6 was found to be elevated in plasma from patients with cervical cancer, with a nearly three-fold increase in patients (966.27 pg/mL in patients vs. 363.54 pg/mL in controls). CMTM6 was preferentially, but not exclusively, found in the exosome-enriched plasma fraction, and was positively correlated with exosomal PD-L1; CMTM6 was identified in the membrane, intracellular compartments, and culture supernatant of the cell lines. These results highlight that CMTM6, in its various presentations, may play an important role in the biology of tumor cells and in immune system evasion.

Pentoxifylline and the proteasome inhibitor MG132 induce apoptosis in human leukemia U937 cells through a decrease in the expression of Bcl-2 and Bcl-XL and phosphorylation of p65.

BACKGROUND: In Oncology, the resistance of the cancerous cells to chemotherapy continues to be the principal limitation. The nuclear factor-kappa B (NF-κB) transcription factor plays an important role in tumor escape and resistance to chemotherapy and this factor regulates several pathways that promote tumor survival including some antiapoptotic proteins such as Bcl-2 and Bcl-XL. In this study, we investigated, in U937 human leukemia cells, the effects of PTX and the MG132 proteasome inhibitor, drugs that can disrupt the NF-κB pathway. For this, we evaluated viability, apoptosis, cell cycle, caspases-3, -8, -9, cytochrome c release, mitochondrial membrane potential loss, p65 phosphorylation, and the modification in the expression of pro- and antiapoptotic genes, and the Bcl-2 and Bcl-XL antiapoptotic proteins. RESULTS: The two drugs affect the viability of the leukemia cells in a time-dependent manner. The greatest percentage of apoptosis was obtained with a combination of the drugs; likewise, PTX and MG132 induce G1 phase cell cycle arrest and cleavage of caspases -3,-8, -9 and cytochrome c release and mitochondrial membrane potential loss in U937 human leukemia cells. In these cells, PTX and the MG132 proteasome inhibitor decrease p65 (NF-κB subunit) phosphorylation and the antiapoptotic proteins Bcl-2 and Bcl-XL. We also observed, with a combination of these drugs overexpression of a group of the proapoptotic genes BAX, DIABLO, and FAS while the genes BCL-XL, MCL-1, survivin, IκB, and P65 were downregulated. CONCLUSIONS: The two drugs used induce apoptosis per se, this cytotoxicity was greater with combination of both drugs. These observations are related with the caspases -9, -3 cleavage and G1 phase cell cycle arrest, and a decrease in p65 phosphorylation and Bcl-2 and Bcl-XL proteins. As well as this combination of drugs promotes the upregulation of the proapoptotic genes and downregulation of antiapoptotic genes. These observations strongly confirm antileukemic potential.

Addition of pentoxifylline to pegylated interferon-alpha-2a and ribavirin improves sustained virological response to chronic hepatitis C virus: a randomized clinical trial.

BACKGROUND AND AIM: The commonly accepted treatment for hepatitis C virus (HCV) infection, pegylated interferon alpha (PEG INF-alpha) and ribavirin, leads to 50-60% of sustained virological response (SVR). On the other hand, pentoxifylline (PTX) possesses antiviral and hepatoprotector properties. AIM: To investigate whether the addition of PTX to conventional hepatitis C treatment increases SVR. MATERIAL AND METHODS: Seventy two patients of both genders were studied in a randomized fashion; the diagnosis of chronic HCV infection was made according to clinical and laboratory criteria and histopathologically classified according to METAVIR scoring system criteria. HCV viral load was tested by PCR, baseline, and after 6 months of treatment, as well as anti-HCV, anti-hepatitis B virus , and anti-human immunodeficiency virus antibodies by enzyme-linked immunosorbent assay. During 48 weeks, control group patients were treated with PEG INF-alpha- 2a plus ribavirin. PTX was administered to Experimental Group patients prior to the treatment. RESULTS: Demographic data were similar in both groups. Experimental- and control-group subjects were at F2 and F3 states according to the METAVIR classification. The most common HCV genotypes were 1a and 1b (39% in the control group in each case, and 42% in the experimental group in each case). At the end of the study, hepatic enzymes and viral load decreased in both groups to similar values. SVR in the experimental group increased significantly (p < 0.05) when compared with standard therapy alone. CONCLUSION: Addition of PTX to conventional chronic hepatitis C treatment may increase the percentage of patients with SVR.

Haplotypes of [-794(CATT)5-8 /-173G>C] MIF gene polymorphisms and its soluble levels in cutaneous squamous cell carcinoma in western Mexican population.

BACKGROUND: Some cytokines are strongly implicated in the development of squamous cell carcinoma (SCC) such as the Macrophage migration inhibitory factor (MIF). The haplotype -794 (CATT)5-8 /-173G>C in MIF gene polymorphisms has been associated with some types of cancer. The aim of this study is to establish the possible association between the presence of this haplotype in the MIF gene and its subsequent soluble levels with the susceptibility of SCC in western Mexican population. METHODS: This study included 175 SCC patients and 175 age-sex-matched individuals as a reference group (RG) from western Mexico. Genomic DNA was extracted from peripheral blood leukocytes. Polymorphisms were genotyped by endpoint PCR and PCR-RFLP, and the determination of MIF serum levels was measured by ELISA. Clinical characteristics were evaluated by a group of dermatologists. RESULTS: Analysis of [-794(CATT)5-8 /-173G>C] MIF gene polymorphisms showed that the 5C (OR = 2.7, p = 0.02) and the 7G (OR = 3.39, p < 0.01) haplotypes are associated with susceptibility in SCC. MIF soluble levels in SCC patients showed a median of 13.93 ng/mL, whereas the reference group showed 6.000 ng/mL. CONCLUSIONS: Our findings suggest that 5C and 7G [-794(CATT)5-8 /-173G>C] MIF gene haplotypes are associated with susceptibility to SCC and that SCC patients present increased soluble levels of MIF.

Pentoxifylline sensitizes human cervical tumor cells to cisplatin-induced apoptosis by suppressing NF-kappa B and decreased cell senescence.

BACKGROUND: Worldwide, cervical cancer is the second most common causes of cancer in women and represents an important mortality rate. Cisplatin (CIS) is a very important antitumoral agent and can lead tumor cells toward two important cellular states: apoptosis and senescence. In some types of cancers pentoxifylline (PTX) sensitizes these cells to the toxic action of chemotherapeutics drugs such as adriamycin, inducing apoptosis. In the present work, we studied in vitro whether PTX alone or in combination with CIS induces apoptosis and/or senescence in cervix cancer HeLa and SiHa cell lines infected with HPV types 16 and 18, respectively, as well as in immortalized keratinocytyes HaCaT cells. METHODS: HeLa (HPV 18+), SiHa (HPV 16+) cervix cancer cells and non-tumorigenic immortalized HaCaT cells (control) were treated with PTX, CIS or both. The cellular toxicity and survival fraction of PTX and CIS were determinate by WST-1 and clonogenic assays respectively. Apoptosis, caspase activation and phosphorylation of ERK1/2, p38, p65 (NF-κB), Bcl-2 and Bcl-XL anti-apoptotic proteins were determinated by flow cytometry. Senescence by microscopy. Phosphorylation of IκBα and IκB total were measured by ELISA. Pro-apoptotic, anti-apoptotic and senescence genes, as well as HPV-E6/7 mRNA expression, were detected by RT-PCR. RESULTS: Our results show that after 24 hours of incubation PTX per se is toxic for cancer cells affecting cell viability and inducing apoptosis. The toxicity in HaCaT cells was minimal. CIS induces apoptosis in HeLa and SiHa cells and its effect was significantly increases when the cells were treated with PTX + CIS. In all studies there was a direct correlation with levels of caspases (-3, -6, -7, -9 and -8) activity and apoptosis. CIS induces important levels of senescence and phosphorylation of ERK1/2, p38, p65/RELA, and IκBα, and decreased the expression of anti-apoptotic protein Bcl-XL. Surprisingly these levels were significantly reduced by PTX in tumor cells, and at the same time, increases the expression of pro-apoptotic genes. CONCLUSION: PTX sensitizes cervical cancer cells to CIS-induced apoptosis and decreases the CIS-induced senescence in these cells via inhibition of NF-κB signaling pathway; diminishes expression of antiapoptotic proteins and the activation of caspases.

Molecular detection of myxoma virus in the environment of vaccinated rabbitries.

Myxoma virus (MYXV) is the aetiological agent of myxomatosis, a systemic, mostly lethal disease that affects European rabbits. Vaccination against it, although widespread, has not been completely effective and disease outbreaks still take place on farms which carry out vaccination programmes. Since some of these cases have been attributed to airborne transmission or the spread of the virus via inanimate vectors, the aims of this study were to determine MYXV contamination levels and distribution in the environment of vaccinated farms and to ascertain whether the detected virus corresponded to field strains. For that, environmental samples from several areas, tools and employees from four (three infected and one uninfected) rabbitries were taken and analysed by qPCR. MYXV was detected in the environment of all the infected farms, whereas all the samples from the non-infected farm were negative. Furthermore, all the positive samples contained viral DNA compatible with field strains of the virus. These results lead us to believe that the administration of currently available commercial vaccines does not prevent infected animals from shedding the field virus. Moreover, viral DNA was also found in items that are not in direct contact with the animals, which could play a role in the transmission of the infection throughout the farm and to other farms. Therefore, this study proves that current vaccination schemes on their own are not sufficient to prevent this disease and should be accompanied by adequate biosecurity measures.

A School-Based Randomized Controlled Trial to Promote Cycling to School in Adolescents: The PACO Study.

This manuscript describes the rationale and protocol of a school-based randomized controlled trial called "Cycling and Walk to School" (PACO, by its Spanish acronym) that aims to promote cycling to and from school and physical activity (PA) in adolescents. This study will examine the effects of this intervention in cycling and active commuting to and from school (ACS), PA and several ACS-related factors based on self-determination theory (SDT) and a social-ecological model (SEM). A total of 360 adolescents attending six high schools (three experimental and three control) from three Spanish cities will participate in this randomized controlled trial. The intervention (four cycling sessions; 1-2 h per session, one session per week) will be conducted by the research staff; the control group will continue their usual activities. PA levels will be measured by accelerometers, whereas ACS and the other study variables will be self-reported using questionnaires at baseline and post-intervention. The primary outcomes will be: rates of cycling to school, ACS and PA levels. In addition, SDT-related variables and individual, interpersonal, community, and environment variables relevant to ACS will be based on SEM. The findings will provide a comprehensive understanding of the short-term effects of this school-based intervention on cycling to school behaviour, ACS and PA levels in Spanish adolescents.

Psychometric Characteristics of a Commuting-to-School Behaviour Questionnaire for Families.

The purposes of this study were: (a) to describe the patterns of modes of commuting to school (children) and to work (parents) separated by gender and age, (b) to validate the questions on children's mode of commuting to and from school according to their parents, and (c) to analyse the reliability of a family questionnaire focused on commuting to school behaviours. A total of 611 parents (mean age: 43.28 ± 6.25 years old) from Granada (Spain) completed "Family commuting-to-school behaviour" questionnaire in two sessions separated by 14 days, (2016 and 2018). The validation between family and children's questions was assessed using the Kappa and Spearman correlation coefficients, and the test-retest reliability within the family questions was assessed using the Kappa and the weighted Kappa. The children's modes of commuting to school (mean age: 11.44 ± 2.77 years old) were mainly passive (57.7% to school) while parents' modes of commuting to work were mainly active (71.6%). The validity of the mode of commuting was significant with high Kappa and Spearman coefficients. The test-retest reliability presented a good agreement for the mode of commuting to school in children, distance and time to school, and the mode of commuting to work in parents, while the questions on acceptable distance to walk or cycle to school showed a moderate to good agreement. The "Family commuting-to-school behaviour" questionnaire could be a useful tool to assess the mode of commuting of children, distance and time to school for researchers and practitioners.

Feasibility and Reliability of a Questionnaire to Assess the Mode, Frequency, Distance and Time of Commuting to and from School: The PACO Study.

Active commuting to and from school has several health implications. Self-reporting is the most common assessment tool, but there is a high heterogeneity of questionnaires in the scientific literature. The purpose of this study was to analyse the feasibility and reliability of the Spanish "New Version of Mode and Frequency of Commuting To and From School" questionnaire in children and adolescents. A total of 635 children (5-12 years old) and 362 adolescents (12-18 years old) filled out the questionnaire twice (14 days apart). Feasibility was evaluated using an observational checklist. The test-retest reliability of the "New Version of Mode and Frequency of Commuting To and From School" questionnaire and the distance and time to school were examined using the kappa and weight kappa coefficient (κ). No misunderstanding of questions was reported. The time to complete the questionnaire was 15 ± 3.62 and 9 ± 2.26 min for children and adolescents, respectively. The questionnaire showed substantial and almost perfect kappa coefficients for the overall six items (k = 0.61-0.94) in children and adolescents. The "New Version of Mode and Frequency of Commuting To and From School" questionnaire is a feasible and reliable questionnaire in Spanish children and adolescents.

Angiofibroma of the Vagina Presenting with Abnormal Vaginal Bleeding: A Case Report from Ethiopia and Review of the Literature.

BACKGROUND: Angiofibroma is an uncommon type of vascular benign tumor that is made up of blood vessels and fibrous (connective) tissue. First described in 1997, it usually occurs in middle aged females and is clinically often thought to represent a cyst. Unlike most of the other site specific vulvovaginal mesenchymal lesions, cellular angiofibroma has a marked predilection for the vulva with only occasional examples reported in the vagina. CASE DETAIL: A 17-year adolescent nulligravid girl presented with a history of irregular vaginal bleeding of two-year duration and history of lower abdominal swelling; on examination, she had pale conjunctiva, 20-week sized firm, irregular, nontender abdominopelvic mass, and a firm huge anterior vaginal wall mass, with difficulty to reach at the cervix and hemoglobin of 9.7 gm/dL, and a diagnosis of cervical myoma plus anemia was made, which was supported by imaging studies. Finally it was found to be angiofibroma of the vagina. CONCLUSION: Angiofibromas are benign tumors, which rarely occur in vagina. Although middle aged females are affected more, angiofibromas can affect females of reproductive age group and can cause abnormal uterine bleeding.

Monitoring of the shedding and serological dynamics of Bovine gammaherpesvirus type 4 in a dairy cattle herd.

Bovine gammaherpesvirus type 4 (BoHV-4) is increasingly related with reproductive disease in cattle, but its epidemiology is not fully understood. We monitored the serological response and shedding of BoHV-4 in a positive dairy cattle farm with metritis. First, we performed an ELISA to detect BoHV-4 antibodies in all the animals (n = 104). Afterwards, ten seronegative heifers introduced in the production lot and sera samples were monthly taken for four months and then 6-10 months after introduction to detect BoHV-4 antibodies by ELISA. Moreover, a vaginal swab was taken after calving to detect BoHV-4 by PCR. Concurrently, a weekly collection of vaginal and nasal swabs and milk was performed during the first month post-partum in multiparous cows with metritis (n = 14), heifers with metritis (n = 4), heifers without metritis but positive to BoHV-4 (ELISA or PCR) (n = 2) and multiparous cows without metritis (n = 3). Seropositivity was higher in older animals and in the production lot. Three heifers which shed BoHV-4 after parturition resulted seronegative at first but eventually seroconverted. In the same vein, most heifers seroconverted after 6-10 months in the production lot (8/10). Multiparous cows shed virus by various routes: 13/14 (93 %) in vaginal secretions, 7/14 (50 %) in nasal exudates and 7/14 (50 %) in milk. However, in the other groups, shedding was only detected in vaginal swabs from the first week post-partum. Our study describes BoHV-4 shedding in field conditions. Seronegative animals may become horizontally infected when moved to a contaminated environment.

An innovative quick solidifying technique for the forensic investigation of brain circulation using addition silicones.

Post-mortem study of the brain is extremely relevant to medico-legal autopsies. However, it can be difficult to handle due to its fragility. This article presents a study on the development of an arterial solidifying technique that can be applied to analyze arterial circulation, consequently easing the handling and later diagnosis of diseases in this anatomical site. Vinylpolysiloxane silicone is introduced into the internal carotid arteries until it completely fills the arterial tree, creating a detailed model of the arterial's internal anatomy. This technique is fast, easy to apply and requires no previous tissue fixation. In addition, it allows for further toxicological and pathological tests. In conclusion, this technique represents a simple, sensitive and efficient method to employ in conventional autopsies, which can help in the diagnosis of death.

Anticipation in familial lattice corneal dystrophy type I with R124C mutation in the TGFBI (BIGH3) gene.

PURPOSE: To report the clinical, ophthalmic, and genetic characteristics for lattice corneal dystrophy type I (LCDI) in a Chilean family. METHODS: Six affected family members were examined clinically including visual acuity, color cornea photography, applanation tonography, and fundoscopy. Genomic DNA was extracted from peripheral leukocytes from six affected and three unaffected members of a family with lattice corneal dystrophy type I. Exon 4 of the transforming growth factor-induced gene (TGFBI) was screened for the most frequent mutation, R124C, in the proband by sequencing. We also designed a rapid polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method to analyze the same mutation, amplifying exon 4 and digesting with PstI restriction enzyme. Using this strategy, we analyzed the mutation in six affected and three healthy family members. RESULTS: Three generations of family members were positively diagnosed with lattice corneal dystrophy. Six participants demonstrated LCD1 in both eyes, most of whom were symmetric. Age at onset of symptoms was variable (3-42 years old). Moreover, in this family, the age of onset of the disease decreased in succeeding generations, which could be interpreted as anticipation. Visual acuity varied from 1.0 to 0.13. Two patients, ages 69 and 44 years old, demonstrated a degree of severity "Bad" according to best-corrected vision and corneal commitment. The exon 4 sequence of TGFBI of the proband exhibits the heterozygous single-nucleotide mutation, C417T, leading to amino acid substitution (R124C) in the encoded TGF-induced protein. Using PCR-RFLP, we confirmed the heterozygous mutation in six affected family members and excluded it in three healthy members. CONCLUSIONS: The R124C mutation in TGFBI cosegregated with LCD type I in the investigated family. This is the first report of a molecular analysis of LCD type I in Chilean patients. The early onset affected persons in the fourth generation raises the possibility of anticipation.