Time-efficient germ cell transplantation from goldfish (Carassius auratus) into adult common carp (Cyprinus carpio).

Germ cell transplantation in fish is a promising technique for surrogate broodstock parents with broader application in aquaculture and conserving endangered and valuable genetic resources. Herein, we describe the establishment of an intrapapillary xenogeneic transplant of germ cells from sexually mature goldfish (C. auratus) males into common carp (C. carpio) males cytoablated with a thermochemical treatment (two doses of busulfan at 40 mg/kg at 35°C). To analyze the presence and development of donor germ cells in recipient testes, donor germ cells were labeled with PKH26, a fluorescent cell membrane dye, before transplantation. Our results demonstrated that thermochemical treatment caused effective spermatogenesis suppression and pronounced germ cell loss. Moreover, transplanted spermatogonial cells were able to colonize the recipients' testes, resume spermatogenesis, and generate spermatozoa within eight weeks after germ cell transplantation. These findings suggested that recipient testes provided suitable conditions for the survival, colonization, proliferation, and differentiation of donor spermatogonia from a related species. This study indicated that recipients' testes exhibited a high degree of plasticity to accept and support xenogeneic donor germ cells, which were able to form sperm in a short time frame. This approach has significant implications for assisted animal reproduction, biotechnology, conservation, and the production of valuable genetic resources and endangered fish species.

A83-01 and DMH1 effects in the zebrafish spermatogonial niche: Unraveling the roles of TGF-ß and BMP signaling in the Fsh-mediated spermatogonial fate.

Transforming growth factor-ß (TGF-ß) and bone morphogenetic protein (BMP) signaling has fundamental roles in the regulation of the stem cell niche for both embryonic and adult stem cells. In zebrafish, male germ stem cell niche is regulated by follicle-stimulating hormone (Fsh) through different members of the TGF-ß superfamily. On the other hand, the specific roles of TGF-ß and BMP signaling pathways are unknown in the zebrafish male germ stem cell niche. Considering this lack of information, the present study aimed to investigate the pharmacological inhibition of TGF-ß (A83-01) and BMP (DMH1) signaling pathways in the presence of recombinant zebrafish Fsh using testicular explants. We also reanalyzed single cell-RNA sequencing (sc-RNA-seq) dataset from adult zebrafish testes to identify the testicular cellular sites of smad expression, and to understand the physiological significance of the changes in smad transcript levels after inhibition of TGF-ß or BMP pathways. Our results showed that A83-01 potentiated the pro-stimulatory effects of Fsh on spermatogonial differentiation leading to an increase in the proportion area occupied by differentiated spermatogonia with concomitant reduction of type A undifferentiated (Aund) spermatogonia. In agreement, expression analysis showed lower mRNA levels for the pluripotency gene pou5f3, and increased expression of dazl (marker of type B spermatogonia and spermatocyte) and igf3 (pro-stimulatory growth factor) following the co-treatment with TGF-ß inhibitor and Fsh. Contrariwise, the inhibition of BMP signaling nullified the pro-stimulatory effects of Fsh, resulting in a reduction of differentiated spermatogonia and increased proportion area occupied by type Aund spermatogonia. Supporting this evidence, BMP signaling inhibition increased the mRNA levels of pluripotency genes nanog and pou5f3, and decreased dazl levels when compared to control. The sc-RNA-seq data unveiled a distinctive pattern of smad expression among testicular cells, primarily observed in spermatogonia (smad 2, 3a, 3b, 8), spermatocytes (smad 2, 3a, 8), Sertoli cells (smad 1, 3a, 3b), and Leydig cells (smad 1, 2). This finding supports the notion that inhibition of TGF-ß and BMP signaling pathways may predominantly impact cellular components within the spermatogonial niche, namely spermatogonia, Sertoli, and Leydig cells. In conclusion, our study demonstrated that TGF-ß and BMP signaling pathways exert antagonistic roles in the zebrafish germ stem cell niche. The members of the TGF-ß subfamily are mainly involved in maintaining the undifferentiated state of spermatogonia, while the BMP subfamily promotes spermatogonial differentiation. Therefore, in the complex regulation of the germ stem cell niche by Fsh, members of the BMP subfamily (pro-differentiation) should be more predominant in the niche than those belonging to the TGF-ß (anti-differentiation). Overall, these findings are not only relevant for understanding the regulation of germ stem cell niche but may also be useful for expanding in vitro the number of undifferentiated spermatogonia more efficiently than using recombinant hormones or growth factors.

Cannabidiol improves Nile tilapia cichlid fish welfare.

Cannabidiol (CBD) is a substance derived from Cannabis sativa, widely studied in medicine for controlling neural diseases in humans. Besides the positive effects on humans, it also presents anxiolytic proprieties and decreases aggressiveness and stress in mammals. Therefore, CBD has the potential to increase welfare in reared animals, as it seems to reduce negative states commonly experienced in artificial environments. Here, we tested the effect of different CBD doses (0, 1, 10 and 20 mg/kg) on aggressiveness, stress and reproductive development of the Nile tilapia (Oreochromis niloticus) a fish reared worldwide for farming and research purposes. CBD mixed with fish food was offered to isolated fish for 5 weeks. The 10 mg/kg dose decreased fish's aggressiveness over time, whereas 20 mg/kg attenuated non-social stress. Both doses decreased the baseline cortisol level of fish and increased the gonadosomatic index. However, CBD 1 and 10 mg/kg doses decreased the spermatozoa number. No CBD dose affected feeding ingestion and growth variables, showing that it is not harmful to meat production amount. Despite the effect on spermatozoa, CBD supplementation exhibits high potential to benefit animals' lives in artificial environments. Therefore, we showed for the first time that CBD could be used as a tool to increase non-mammal welfare, presenting a great potential to be explored in other husbandry and captivity species.

In silico Study of Acetylcholinesterase and Beta-secretase Inhibitors: Potential Multitarget Anti-Alzheimer's Agents.

BACKGROUND: Alzheimer's disease is a progressive neurodegenerative process with multifactorial characteristics. This disease follows the natural aging process, affecting mainly people over 65 years. Pharmacotherapeutic treatment currently combats symptoms related to cognitive function. Several targets have begun to attract the interest of the scientific community to develop new drug candidates which have better pharmacokinetic and lower toxicity parameters. OBJECTIVE: The present study aims to design new candidates for acetylcholinesterase/ß-secretase (AChE/BACE1) multitarget inhibitor drugs. METHODS: 17 natural products were selected from the literature with anticholinesterase activity and 1 synthetic molecule with inhibitory activity for BACE1. Subsequently, the molecular docking study was performed, followed by the derivation of the pharmacophoric pattern and prediction of pharmacokinetic and toxicological properties. Finally, the hybrid prototype was designed. RESULTS: All selected molecules showed interactions with their respective target enzymes. Derivation of the pharmacophoric pattern from molecules that interacted with the AChE enzyme resulted in 3 pharmacophoric regions: an aromatic ring, an electron-acceptor region and a hydrophobic region. The molecules showed good pharmacokinetic and toxicological results, showing no warnings of mutagenicity and/or carcinogenicity. After the hybridization process, three hybrid molecules were obtained, which showed inhibitory activity for both targets. CONCLUSION: It is concluded that research in the field of medicinal chemistry is advancing towards the discovery of new drug candidates that bring a better quality of life to patients with AD.

Epigenetic priming by EHMT1/EHMT2 in acute lymphoblastic leukemia induces TP53 and TP73 overexpression and promotes cell death.

Euchromatic histone-lysine N-methyltransferase 1 (EHMT1) and EHMT2 are upregulated in various human cancers, and their deregulation is associated with tumor development and progression. In this paper, we investigated the expression level of EHMT1/EHMT2 in acute lymphoblastic leukemia (ALL) and whether the modulation of these enzymes could have any cellular or molecular impact on ALL cells. For this, we used UNC0646 as a priming strategy to target EHMT1/EHMT2 and investigated its effect on proliferation and cell viability of Jurkat cells by MTT assay. Then, considering the IC50 and IC75, cellular death was determined by Annexin V/PI staining using flow cytometry. Finally, we investigated by RT-PCR the molecular bases that could be involved in the observed effects. Interestingly, accessing the International Microarray Innovations in Leukemia (MILE) study group, we detected that both EHMT1 and EHMT2 are overexpressed in ALL. More important, we determined that inhibition of EHMT1/EHMT2 significantly decreased Jurkat cell viability in a dose-dependent manner. Accordingly, we observed that inhibition of EHMT1/EHMT2 promoted Jurkat cell death, which was accompanied by increased expression of P53, TP73, BAX, and MDM4. These results clearly indicate that inhibition of EHMT1/EHMT2 induces pro-apoptotic gene expression in ALL and promotes cell death. More importantly, the modulation of these histone methyltransferases may be a promising epigenetic target for ALL treatment.

[Helicobacter pylori infection detected during upper gastrointestinal endoscopy]. / Disminución en la frecuencia de infección por Helicobacter pylori en endoscopías digestivas altas.

BACKGROUND: Helicobacter pylori (HP) is the most widespread chronic human infection worldwide and the most important pathogenic factor of gastric cancer. The calculated prevalence at the Clinical Hospital of the University of Chile from 2002 to 2005 was 44.9%. AIM: To determine the current prevalence of HP in patients undergoing an upper gastrointestinal endoscopy (UGI) and analyze its distribution according to age and endoscopic findings. MATERIAL AND METHODS: We reviewed 3.433 UGI performed during the year 2015, selecting those in which rapid urease test (RUT) was done. A positive RUT or a positive gastric biopsy (GB) were considered as HP infection. RESULTS: RUT was done in 1862 UGI (55%) performed in patients aged 51 ± 17 years, (66% women). In 23% of these endoscopies, the RUT was positive. A GB was obtained 43% of endoscopies and 30% were positive for HP. In 105 patients the RUT was negative and the GB positive (rendering a 19.5% false negative rate). HP was detected by RUT and GB in 29% of endoscopies. The highest prevalence of infection (38.1%) was found between 40 and 49 years. HP infection had odds ratio of 4.24 for nodular gastropathy, 2.63 for gastric ulcer and 2.14 for duodenal ulcer (p < 0.05). CONCLUSIONS: HP prevalence in our center decreased significantly from 44.9% to 28.9% in 11 years. False negative RUT results may bias this finding. The use of proton pump inhibitors and antimicrobials that can interfere with the detection of HP should be registered to properly analyze the results of the RUT.

Disminución en la frecuencia de infección por Helicobacter pylori en endoscopías digestivas altas / Helicobacter pylori infection detected during upper gastrointestinal endoscopy

Background: Helicobacter pylori (HP) is the most widespread chronic human infection worldwide and the most important pathogenic factor of gastric cancer. The calculated prevalence at the Clinical Hospital of the University of Chile from 2002 to 2005 was 44.9%. Aim: To determine the current prevalence of HP in patients undergoing an upper gastrointestinal endoscopy (UGI) and analyze its distribution according to age and endoscopic findings. Material and Methods: We reviewed 3.433 UGI performed during the year 2015, selecting those in which rapid urease test (RUT) was done. A positive RUT or a positive gastric biopsy (GB) were considered as HP infection. Results: RUT was done in 1862 UGI (55%) performed in patients aged 51 ± 17 years, (66% women). In 23% of these endoscopies, the RUT was positive. A GB was obtained 43% of endoscopies and 30% were positive for HP. In 105 patients the RUT was negative and the GB positive (rendering a 19.5% false negative rate). HP was detected by RUT and GB in 29% of endoscopies. The highest prevalence of infection (38.1%) was found between 40 and 49 years. HP infection had odds ratio of 4.24 for nodular gastropathy, 2.63 for gastric ulcer and 2.14 for duodenal ulcer (p < 0.05). Conclusions: HP prevalence in our center decreased significantly from 44.9% to 28.9% in 11 years. False negative RUT results may bias this finding. The use of proton pump inhibitors and antimicrobials that can interfere with the detection of HP should be registered to properly analyze the results of the RUT.

Factors Associated with Clinical and Topographical Features of Laryngeal Tuberculosis.

INTRODUCTION: Laryngeal tuberculosis (LTB) is the most frequent granulomatous disease of the larynx and represents less than 2% of extrapulmonary TB cases. There are no pathognomonic clinical and endoscopic features of this disease and studies on LTB that can assist in its diagnostic characterization are lacking. OBJECTIVE: To identify factors associated with clinical and topographical features of LTB. METHOD: a retrospective cross-sectional study was conducted from the medical records of 36 patients with confirmed LTB diagnosis. RESULTS: Dysphonia and cough were the main symptoms presented by patients and the true vocal folds the most frequently affected site. The average of the duration of the disease evolution was significantly higher in patients with dysphonia than in patients without this symptom. We observed association between dysphonia and true vocal fold lesions and between odynophagia and lesions in the epiglottis, arytenoids and aryepiglottic folds. Odynophagia was more frequent in individuals with lesions in four or more laryngeal sites. Weight loss equal or above 10% of the body weight was more frequent in patients with odynophagia as first symptom and in patients with ulcerated lesion. Dyspnea on exertion was more frequent in individuals with more extensive laryngeal lesions. The percentage of smokers with lesions in four or more laryngeal sites was greater than that found in non-smokers. Laryngeal tissue fragment bacilloscopy and culture examinations were less positive than sputum ones. CONCLUSIONS: Smoking appears to be associated with the development of more extensive LTB lesions, and LTB with dyspnea on exertion and odynophagia with consequent impairment of nutritional status. We emphasize the need for histopathologic confirmation, once positive sputum bacteriological examinations seem not to necessarily reflect laryngeal involvement.

Evaluation of voice disorders in patients with active laryngeal tuberculosis.

INTRODUCTION: Laryngeal tuberculosis (LTB) is the most frequent larynx granulomatous disease. In general there is lung involvement, but in an important proportion of cases you can find LTB without pulmonary disease. The lesions observed in LTB, such as ulceration and fibrosis, can interfere in the process of voice production. The involvement of the mucous lining of the vocal folds can change their flexibility and, consequently, change voice quality, and the main symptom is dysphonia present in almost 90% of cases. OBJECTIVE: To describe the anatomical characteristics and voice quality in LTB patients. MATERIAL AND METHOD: A descriptive cross-sectional study was conducted with 24 patients. RESULT: The most frequently affected sites were vocal folds in 87.5% patients, vestibular folds in 66.7%, epiglottis in 41.7%, arytenoid in 50%, aryepiglottic folds in 33.3%, and interarytenoid region in 33.3% patients. We found 95.8% cases of dysphonia. The voice acoustic analysis showed 58.3% cases of Jitter alterations, 83.3% of Shimmer and 70.8% of GNE. CONCLUSION: Voice disorders found in active laryngeal tuberculosis are similar to those reported after clinical healing of the disease, suggesting that sequelae and vocal adjustments may install during the active phase of the disease, negatively impacting the process of vocal quality reestablishment.

Oral manifestations in the American tegumentary leishmaniasis.

INTRODUCTION: American tegumentary leishmaniasis (ATL) can affect the skin or mucosa (mucocutaneous leishmaniasis - MCL) including the oral cavity. MCL oral lesions are often confused with other oral diseases, delaying diagnosis and specific treatment, and increasing the likelihood of sequelae. Thus, increasing the knowledge of the evolution of ATL oral lesions can facilitate its early diagnosis improving the prognosis of healing. OBJECTIVES: Evaluate the frequency of ATL oral lesion and describe its clinical, laboratory and therapeutic peculiarities. METHODS: A descriptive transversal study was carried out, using data from medical records of 206 patients with MCL examined at the outpatient clinics-IPEC-Fiocruz between 1989 and 2013. Proportions were calculated for the categorical variables and the association among them was assessed by the Pearson's chi-square test. Measures of central tendency and dispersion were used for the continuous variables and their differences were assessed by both parametric (t test) and non parametric (Mann-Whitney) tests. P-values <0.05 were considered as significant. RESULTS: The most affected site was the nose, followed by the mouth, pharynx and larynx. Seventy eight (37.9%) have oral lesions and the disease presented a lower median of the evolution time than in other mucous sites as well as an increased time to heal. The presence of oral lesion was associated with: the presence of lesions in the other three mucosal sites; a smaller median of the leishmanin skin test values; a longer healing time of the mucosal lesions; a higher recurrence frequency; and a smaller frequency of treatment finishing and healing. When the oral lesion was isolated, it was associated with an age 20 years lower than when the oral lesion was associated with other mucosal sites. CONCLUSION: Considering the worst therapy results associated with the presence of oral lesions, we suggest that lesions in this location represent a factor of worse prognosis for MCL.

Electrochemical study of ricin at glassy carbon electrode.

Ricin, Ricinus communis agglutinin 60 - RCA 60, is a deadly phytotoxic protein which inhibits ribosomes (class II), and there is no known effective antidote in living organisms. Ricin is composed of two polypeptide chains, A and B, linked covalently by a single disulfide bond. The analytical methods for the detection of RCA 60 are commonly laborious, expensive, require skilled labor, and involve sophisticated equipment. Aimed at the development of electroanalytical methods for RCA 60 detection, here we studied the electrochemical oxidation of RCA 60 on a glassy carbon (GC) electrode over a wide pH range, using cyclic voltammetry, differential pulse voltammetry (DPV) and square wave voltammetry (SWV). Two quasi-reversible electrochemical RCA 60 oxidation peaks were identified on the GC electrode by SWV. For values of 2.2 ≤ pH ≤ 10.2, DPV studies revealed that the peak potentials, EP1 and EP2, display a linear dependence with pH and the reaction mechanism involves the transfer of 2H⁺/2e⁻ (peak 1) and 1H⁺/1e⁻ (peak 2). The first and second RCA 60 oxidation steps may correspond to the oxidation of cysteine and tyrosine-tryptophan residues, respectively. The oxidation product of the second RCA 60 oxidation step appears at 7.0 ≤ pH ≤ 11.8. For pH ≥ 10.2, both processes are pH independent, resulting in a pKa of ca. 10.2. A third RCA 60 oxidation peak only appears at acidic pH. RCA 60 samples extracted from different castor seed cultivars showed similar electrochemical behavior, enabling the implementation of an analytical voltammetric method.

2,2'-Dihydroxy-N,N'-(ethane-1,2-di-yl)di-benzamide.

The asymmetric unit of the title compound, C(16)H(16)N(2)O(4), contains one half-mol-ecule, the whole mol-ecule being generated by an inversion center located at the mid-point of the C-C bond of the central ethane group. An intra-molecular O-H⋯O hydrogen bond forms an S(6) ring motif. In the crystal, mol-ecules are connected via N-H⋯O hydrogen bonds, generating infinite chains along [1-10].

[Cardiogenic shock secondary to acute myocardial infarction managed with high volume hemofiltration: report of a case]. / Shock cardiogénico por infarto agudo del miocardio manejado con hemofiltración de alto volumen: Caso clínico.

Cardiogenic shock secondary to acute myocardial infarction unveils a systemic inflammatory response with elevation of cytokines that contribute to hypoperfusion. High volume hemofiltration may remove cytokines in patients with septic shock resulting in hemodynamic improvement and reducing the requirements of norepinephrine. We report a 48 year-old male with cardiogenic shock secondary to acute myocardial infarction who presented a systemic inflammatory response characterized by fever and hemodynamic collapse, without evidence of infection. Its hemodynamic profile was stabilized with high volume hemofiltration.