Morphobiometry of the scleral ossicle rings in Chelonia mydas sea turtles.

OBJECTIVE: Scleral ossicle rings of reptiles have endoskeletal functions that are not completely understood. Moreover, descriptive reports on the anatomy of those rings are scarce. We tried to make an anatomical description that could contribute to a better understanding of their functions. ANIMAL STUDIED AND PROCEDURES: We quantified, histologically characterized and evaluated the morphobiometry of the scleral ossicles, and measured the aditus orbitae of 25 sea turtle (Chelonia mydas) heads. RESULTS: The aditus orbitae represented about one-third of the total head length and the mean area of the internal opening of each ring was up to 8.37% of the aditus orbitae area. The mean internal diameter of the rings (6.32 mm) was characteristic of scotopic species and the most frequent number of ossicles per ring varied between 11 and 12. Two new classifications were proposed for the ossicle types: plus-Verzahnung (+V) and minus-Verzahnung (-V). The bone tissue revealed a lamellar arrangement typical of compact and resistant bones. CONCLUSION: The obtained data may be used to support and expand the understanding of functions, animal activity patterns, distinctions between taxa and taphonomic interpretations.

H4K20me1 and H3K27me3 are concurrently loaded onto the inactive X chromosome but dispensable for inducing gene silencing.

During X chromosome inactivation (XCI), in female placental mammals, gene silencing is initiated by the Xist long non-coding RNA. Xist accumulation at the X leads to enrichment of specific chromatin marks, including PRC2-dependent H3K27me3 and SETD8-dependent H4K20me1. However, the dynamics of this process in relation to Xist RNA accumulation remains unknown as is the involvement of H4K20me1 in initiating gene silencing. To follow XCI dynamics in living cells, we developed a genetically encoded, H3K27me3-specific intracellular antibody or H3K27me3-mintbody. By combining live-cell imaging of H3K27me3, H4K20me1, the X chromosome and Xist RNA, with ChIP-seq analysis we uncover concurrent accumulation of both marks during XCI, albeit with distinct genomic distributions. Furthermore, using a Xist B and C repeat mutant, which still shows gene silencing on the X but not H3K27me3 deposition, we also find a complete lack of H4K20me1 enrichment. This demonstrates that H4K20me1 is dispensable for the initiation of gene silencing, although it may have a role in the chromatin compaction that characterises facultative heterochromatin.

An eye on the Shih Tzu dog: Ophthalmic examination findings and ocular surface diagnostics.

OBJECTIVE: To characterize the ocular surface parameters and determine the prevalence of ocular pathology in Shih Tzu dogs. ANIMAL STUDIED: Fifty Shih Tzu dogs (28 male, 22 female). PROCEDURES: Each dog underwent a complete ophthalmic examination (recording any pathology) and a series of diagnostics, allowing for a 10 min-interval between tests: intraocular pressure (IOP), blink rate, palpebral fissure length (PFL), corneal tactile sensation (CTS), Schirmer tear test and nasolacrimal reflex without (STT-1, NL-STT1) and with topical anesthesia (STT-2, NL-STT2), tear ferning, strip meniscometry test (SMT), tear film breakup time (TFBUT), and punctate fluorescein staining (PFS) of the cornea. RESULTS: Mean ± SD test values were as follows: IOP (17.9 ± 3.7 mmHg), blink rate (2.4 ± 1.4 blinks/min), PFL (23.8 ± 1.8 mm), CTS (1.8 ± 0.7 cm), STT-1 (22.0 ± 5.5 mm/min), NL-STT1 (24.2 ± 4.7 mm/min), STT-2 (16.9 ± 6.5 mm/min), NL-STT2 (18.5 ± 7.5 mm/min), SMT (7.5 ± 3.5 mm/5 s), TFBUT (5.3 ± 2.4 s), tear ferning (1.3 ± 0.7), and PFS (1.6 ± 0.6). PFL was significantly greater in male vs. female Shih Tzus (p< .001). Age was negatively correlated with TFBUT results (r = -0.31, p = .027). Lagophthalmos was observed in 82% eyes. Ocular surface pathology was common, including adnexal abnormalities (100% eyes with caruncular trichiasis and medial lower lid entropion) and corneal opacification (27% pigmentation, 20% fibrosis, 12% neovascularization). CONCLUSIONS: Qualitative tear film deficiency (low TFBUT), along with several anatomical abnormalities that promote ocular irritation and reduce globe protection, together help explain the concerningly high prevalence of ocular surface disease in the Shih Tzu breed. Prophylactic measures (e.g., medial canthoplasty, topical lubrication) could be considered to improve ocular health in Shih Tzus.

TonoVet Plus®: Higher reliability and repeatability compared with Tono-Pen XL™ and TonoVet® in rabbits.

OBJECTIVE: To evaluate the accuracy and repeatability of the Tono-Pen XL™, TonoVet® and TonoVet Plus® tonometers by manometric evaluation, and to establish adjustment equations for intraocular pressure (IOP) estimates in rabbits. ANIMAL STUDIED: Rabbits. PROCEDURES: A postmortem study was conducted on seven rabbit eyes to verify the correlation between manometry and tonometry with an artificial incremental increase in IOP from 5 and 60 mmHg. A clinical study was conducted to evaluate accuracy and to establish reference values for the species, with measurement of IOP in 17 animals, for 2 consecutive days, with the same tonometers and calibrations used in the postmortem evaluations. RESULTS: There were strong linear trends for all evaluated tonometers. In the in-vivo evaluation, the mean IOP values were: 14.23 ± 1.75 (Tono-Pen XL™); 13.89 ± 2.07 (TonoVet® calibration mode 'd'); 8.88 ± 1.24 (TonoVet calibration mode 'p'); 18.59 ± 1.94 (Tonovet Plus®). There was a significant difference in the two evaluation times for the two TonoVet® calibration modes. The adjustment equations generated from the manometry for the evaluated tonometers were: Y = 0.2570X + 2.219 (Tono-Pen XL™), Y = 0.2289X + 2.389 (TonoVet® 'd'), Y = 0.4043X + 4.062 (TonoVet® 'p'), Y = 0.1233X + 0.3644 (TonoVet Plus®) (X is device-estimated IOP). CONCLUSIONS: All evaluated tonometers were well correlated with the manometry, with an underestimation of IOP by all devices. Applying adjustment formulas may compensate for systematic errors. TonoVet Plus® was well tolerated, and showed better repeatability and reliability in successive evaluations.

The tandem repeat modules of Xist lncRNA: a swiss army knife for the control of X-chromosome inactivation.

X-inactive-specific transcript (Xist) is a long non-coding RNA (lncRNA) essential for X-chromosome inactivation (XCI) in female placental mammals. Thirty years after its discovery, it is still puzzling how this lncRNA triggers major structural and transcriptional changes leading to the stable silencing of an entire chromosome. Recently, a series of studies in mouse cells have uncovered domains of functional specialization within Xist mapping to conserved tandem repeat regions, known as Repeats A-to-F. These functional domains interact with various RNA binding proteins (RBPs) and fold into distinct RNA structures to execute specific tasks in a synergistic and coordinated manner during the inactivation process. This modular organization of Xist is mostly conserved in humans, but recent data point towards differences regarding functional specialization of the tandem repeats between the two species. In this review, we summarize the recent progress on understanding the role of Xist repetitive blocks and their involvement in the molecular mechanisms underlying XCI. We also discuss these findings in the light of the similarities and differences between mouse and human Xist.

The role of Xist-mediated Polycomb recruitment in the initiation of X-chromosome inactivation.

Xist RNA has been established as the master regulator of X-chromosome inactivation (XCI) in female eutherian mammals, but its mechanism of action remains unclear. By creating novel Xist-inducible mutants at the endogenous locus in male mouse embryonic stem (ES) cells, we dissect the role of the conserved A-B-C-F repeats in the initiation of XCI. We find that transcriptional silencing can be largely uncoupled from Polycomb repressive complex 1 and complex 2 (PRC1/2) recruitment, which requires B and C repeats. Xist ΔB+C RNA specifically loses interaction with PCGF3/5 subunits of PRC1, while binding of other Xist partners is largely unaffected. However, a slight relaxation of transcriptional silencing in Xist ΔB+C indicates a role for PRC1/2 proteins in early stabilization of gene repression. Distinct modules within the Xist RNA are therefore involved in the convergence of independent chromatin modification and gene repression pathways. In this context, Polycomb recruitment seems to be of moderate relevance in the initiation of silencing.

The glycoproteomics of hawk and caiman tears.

BACKGROUND: Glycoproteins are important tear components that participate in the stability of the ocular surface. However, the glycopeptides that are present in the tears of wild animals have not yet been described. This work aimed to describe the glycoproteomic profile of roadside hawk (Rupornis magnirostris) and caiman (Caiman latirostris) tears. METHODS: Tears collected from 10 hawks and 70 caimans using Schirmer tear test strips were used in this study. The samples were submitted to trypsin digestion and separated using a reverse-phase column coupled to a mass spectrometer associated to a nanospray ionization source. The glycoproteins were categorized as: cellular components, biological processes and molecular function, according to the UniProt Knowledgebase. RESULTS: As shown by the liquid chromatography-mass spectrometry, all glycopeptides found were classified as N-type. Of the 51 glycoproteins that were identified in the hawk tear film, the most abundant were ovotransferrin, globulins and complement system proteins. In the caiman tear film, 29 glycoproteins were identified. The most abundant caiman glycoproteins were uncharacterized proteins, ATPases, globulins and proteasome components. Ontological characterization revealed that the glycoproteins were extracellular, and the most identified molecular function was endopeptidase activity for both species. CONCLUSION: Glycoproteins are abundant in the tear film of the bird and reptile species studied herein, and all these molecules were shown to have N-type modifications. Location at the extracellular space and an endopeptidase inhibitor activity were the main cell component and molecular function for both species, respectively. These profiles showed differences when compared to human tears, are possibly linked to adaptive processes and can be the basis for further studies on the search of disease biomarkers.

Atypical ocular Chelonoid herpesvirus manifestations in a captive Loggerhead turtle (Caretta caretta).

A captive loggerhead turtle (Caretta caretta) of unknown sex, 3 years of age, presented with bilateral mucoid secretions, severe chemosis, conjunctival hyperemia, and globe retraction. The animal was evaluated ophthalmologically and systemically, and hematological, microbiological, and conjunctival cytological and biopsy samples were collected for complementary diagnosis. The histopathological examination showed amphophilic intranuclear inclusions associated with severe inflammatory infiltrate. The diagnosis of Chelonid alphaherpesvirus 5 (ChAHV 5) was confirmed with end point PCR. Following systemic treatment with L-lysine, acyclovir and vitamin A, the ocular signs resolved. No amphophilic intranuclear inclusions were seen in a follow-up biopsy 5 months later, and there has been no recurrence of clinical ophthalmic signs during a 4-year follow-up. It is suggested that ChAHV 5 be considered as a differential diagnosis in captive marine turtles that present for conjunctival disease other than fibropapillomatosis.

Loss of hierarchical imprinting regulation at the Prader-Willi/Angelman syndrome locus in human iPSCs.

The human chr15q11-q13 imprinted cluster is linked to several disorders, including Prader-Willi (PWS) and Angelman (AS) syndromes. Recently, disease modeling approaches based on induced pluripotent stem cells (iPSCs) have been used to study these syndromes. A concern regarding the use of these cells for imprinted disease modeling is the numerous imprinting defects found in many iPSCs. Here, by reprogramming skin fibroblasts from a control and AS individuals, we generated several iPSC lines and addressed the stability of imprinting status across the PWS/AS domain. We focused on three important regulatory DNA elements which are all differentially methylated regions (DMRs), methylated on the maternal allele: the PWS imprinting center (PWS-IC), which is a germline DMR and the somatic NDN and MKRN3 DMRs, hierarchically controlled by PWS-IC. Normal PWS-IC methylation pattern was maintained in most iPSC lines; however, loss of maternal methylation in one out of five control iPSC lines resulted in a monoallelic to biallelic switch for many imprinted genes in this domain. Surprisingly, MKRN3 DMR was found aberrantly hypermethylated in all control and AS iPSCs, regardless of the methylation status of the PWS-IC master regulator. This suggests a loss of hierarchical control of imprinting at PWS/AS region. We confirmed these results in established iPSC lines derived using different reprogramming procedures. Overall, we show that hierarchy of imprinting control in donor cells might not apply to iPSCs, accounting for their spectrum of imprinting alterations. Such differences in imprinting regulation should be taken into consideration for the use of iPSCs in disease modeling.

Aqueous tear assessment in dogs: Impact of cephalic conformation, inter-test correlations, and test-retest repeatability.

OBJECTIVE: To characterize diagnostic findings, test-retest repeatability, and correlations among lacrimal tests in dogs of diverse cephalic conformations. ANIMAL STUDIED: Fifty healthy dogs (25 brachycephalic, 25 nonbrachycephalic). PROCEDURES: A series of diagnostics were performed in each dog, allowing for a 10-minute interval between tests and repeating each test 24 hours later under similar conditions: corneal tactile sensation (CTS), strip meniscometry test (SMT), phenol red thread test (PRTT), endodontic absorbent paper point tear test (EAPPTT), Schirmer tear test-1 without (STT-1) or with nasolacrimal stimulation (NL-STT1), and Schirmer tear test-2 (STT-2). RESULTS: Mean ± SD test values were lower in brachycephalic vs. nonbrachycephalic dogs (except for SMT; 7.4 ± 2.0 mm/5 seconds vs 7.3 ± 2.4 mm/5 seconds), with statistically significant differences noted for CTS (1.8 ± 0.5 cm vs 3.4 ± 0.8 cm), PRTT (37.2 ± 4.0 mm/15 seconds vs 41.1 ± 5.5 mm/15 seconds), STT-1 (20.1 ± 3.4 mm/min vs 23.3 ± 5.7 mm/min), STT-2 (13.0 ± 3.4 mm/min vs 16.9 ± 3.9 mm/min), and NL-STT1 (23.2 ± 3.6 mm/min vs 27.1 ± 5.4 mm/min), and nonsignificant differences for EAPPTT (16.6 ± 2.7 mm/15 seconds vs 17.5 ± 2.9 mm/15 seconds). Nasolacrimal stimulation increased STT-1 values by 18% on average. Correlations among tests were generally weak to moderate (r < .70) except for a strong correlation between STT-1 and NL-STT1 (r = .83, P < .001). Test reliability was good although test-retest repeatability was generally poor to moderate, as depicted by low intraclass correlation coefficients (ICC ≤ 0.75) and wide 95% limits of agreement, except for CTS (ICC = 0.91). CONCLUSIONS: Corneal sensitivity and aqueous tear secretion are lower in brachycephalic dogs. A comprehensive assessment of the ocular surface requires the combination of several diagnostic tests. The nasolacrimal reflex may provide a useful diagnostic and therapeutic tool in dogs.

Conjunctival bacterial flora and antimicrobial susceptibility of captive and free-living sea turtles in Brazil.

PURPOSE: To describe the aerobic conjunctival bacterial flora of 3 especies of free-living and under human care sea turtles and determine its antimicrobial susceptibility in vitro. METHOD: Thirty-six sea turtles (72 eyes), juveniles and adults, 7 free-living Chelonia mydas and 8 Chelonia mydas, 4 Caretta caretta, 11 Eretmochelys imbricata, and 6 Lepidochelys olivacea under human care, were evaluated. Conjunctival cultures were collected for identification of aerobic bacteria and antimicrobial susceptibility testing for ciprofloxacin, chloramphenicol, gentamicin, neomycin, oxacillin, polymyxin B, tetracycline, and tobramycin using antibiotic disks. Bacterial strains showing no sensitivity to 4 or more antimicrobials were considered multiresistant to this panel. RESULTS: Bacterial growth was observed in 12/14 (85.71%) samples in the free-living sea turtles, and there was growth in 100% (58/58) of the samples from captive animals. There were 94 strains isolated and 15 species identified. There was a predominance of Gram-positive bacteria in free-living Chelonia mydas, most of which were Bacillus and Staphylococcus. The most commonly isolated Gram-negative species were enterobacteria for free-living and under human care animals. The strains were predominantly sensitive to ciprofloxacin and tobramycin, and less sensitive to oxacillin or polymyxin B. Ten multiresistant strains were isolated. Yeast were identified in 13.89% (10/72) of the samples. CONCLUSIONS: These results, showing differences in the conjunctival bacterial flora of free-living and captive animals, may be helpful for diagnosis and treatment of ocular disorders in sea turtles.

Evaluation of lacrimal production, osmolarity, crystallization, proteomic profile, and biochemistry of capuchin monkeys' tear film.

BACKGROUND: To evaluate the composition and characteristics of capuchin monkey (CM) tear film. METHODS: Eleven CM (Sapajus sp.) was evaluated. Strip meniscometry test (SMT), osmolarity, and tear ferning test (TFT) (by Rolando and Masmali scales) were assessed. Tear protein profile (SDS-PAGE), and total protein, albumin, urea, glucose, and cholesterol levels in tear film and blood serum were screened. RESULTS: Median ± semi-interquatil range for SMT and osmolarity values were 8.0 ± 1.625 and 303.0 ± 9.875, respectively. TFT for Rolando was 2.0 ± 0.5, and Masmali was 2.0 ± 0.0. Monkeys's tear obtained type II and III for Rolando, and 1 and 2 for Masmali. Tear components showed protein bands among 23-217 kDa, and presence of albumin, urea, glucose, and cholesterol. CONCLUSIONS: The results of SMT, osmolarity, TFT, SDS-PAGE, and tear biochemistry may serve as a reference baseline for CM, and the data may serve as a basis for future experimental model evaluations.

Correction to: Photography-based method for assessing fluorescein clearance test in dogs.

The original article [1] contained an error whereby the respective legends of Figs. 2 and 3 were mistakenly interchanged. This error has now been amended.

Photography-based method for assessing fluorescein clearance test in dogs.

BACKGROUND: The fluorescein clearance test (FCT) provides insight into the tear film dynamics. The purpose of this study was to describe an inexpensive and practical method for assessing FCT in dogs, using photography and software analysis, and to assess the retention time of 1 vs. 2 eye drops on the canine ocular surface. METHODS: (i) In vivo - Eight healthy German Shepherd dogs were recruited. Following topical anesthesia with 0.5% proxymetacaine, each eye sequentially received (1 week apart) either 1 drop (35 µL) or 2 drops (70 µL) of 0.5% fluorescein. A Schirmer strip was inserted in the ventral conjunctival fornix for 10 s at the following times: each 10 min for 100 min, 24 h, 48 h and 72 h. (ii) In vitro - Schirmer strips were placed for 10 s in contact with microplate wells containing 1 or 2 drops of 0.5% fluorescein. In both experiments, the fluorescein-impregnated Schirmer strips were immediately imaged, and the area and intensity of fluorescein uptake were analyzed with ImageJ software. For the in vitro experiment, images were evaluated by the same examiner (repeatability) or two examiners (reproducibility). RESULTS: Photography-based FCT was easy to perform and showed high repeatability and reproducibility (coefficients of variation ≤2.75%). In vivo, the area and intensity of fluorescein uptake on Schirmer strips were significantly greater at 30 min and 40 min post- fluorescein instillation in the 2 drops vs. 1 drop groups (p ≤ 0.044). Compared to baseline, the residual fluorescein uptake on Schirmer strips was < 5% at 60 min and 90 min in the 1 drop and 2 drops groups, respectively. CONCLUSIONS: Photography-based FCT is a practical and reliable diagnostic tool with various clinical and research applications in veterinary medicine. Instillation of two drops provided greater amount and longer retention on the anesthetized canine ocular surface than a single drop. Fluorescein clearance time of a single drop in dolichocephalic dogs is 60 min.

Tear ferning test in healthy dogs.

PURPOSE: To evaluate and compare three tear sampling methods using two grading scales for administering the tear ferning test (TFT) to healthy dogs. METHODS: In total, 90 dogs (180 eyes) were subjected to tear sampling using millimetered strips, reused after the Schirmer tear test (STT) (Schirmer group, SG). Then, the dogs were subdivided into three groups according to sampling approach: micropipette (MPG), microcapillary (MCG), and Schirmer sample 2 (S2G). The collected tears were dried on a clean microscope glass slide at room temperature and humidity. The ferning patterns were observed under a polarized light microscope and classified according to the Rolando and Masmali grading scales. RESULTS: Although all three methods were feasible, the STT was easier to perform in clinical settings. Type I and Grade 1 were the most commonly observed (64.17% and 61.7%, respectively) regardless of collection method. There was no significant difference between the STT median values and the TFT classifications. CONCLUSIONS: The TFT is appropriate for dogs and can be performed using the three suggested sampling methods, with a higher frequency of Type I and Grade 1. Thus, it is possible to use both grading scales in the classification of tear ferning in dogs.

CONJUNCTIVAL BACTERIAL FLORA, ANTIBIOGRAM, AND LACRIMAL PRODUCTION TESTS OF COLLARED ANTEATER (TAMANDUA TETRADACTYLA).

The collared anteater ( Tamandua tetradactyla ) is adapted to a variety of habitats. It is a solitary species for which no reference values for ophthalmic tests have been established. Eight animals ranging from 1 to 4 yr of age, two males and six females, were manually restrained for assessment. Ophthalmic tests included evaluation of tear production by Schirmer tear test 1 (STT1), endodontic absorbent paper point tear test (EAPPTT), palpebral fissure length (PFL), culture of the conjunctival bacterial flora, and antimicrobial susceptibility test. Median ± semi-interquartile range (S-IQR) STT1, EAPPTT, and PFL were 8.50 ± 4.13 mm/min, 14.13 ± 3.24 mm/min, and 15.91 ± 2.51 mm, respectively. Bacterial growth was present in 100% of the samples, with predominance of Gram-positive bacteria (70.27%). Staphylococcus spp. was the most frequently isolated genus. Antimicrobial susceptibility test showed sensitivity of Staphylococcus epidermidis and Staphylococcus spp. to neomycin, tobramycin, and gentamicin. The results in this study can benefit the determination of reference values for different diagnostic techniques, and may be used as a guide for diagnosis and treatment of ocular diseases in collared anteaters.

Sedative Effects of Intranasal Midazolam Administration in Wild Caught Blue-fronted Amazon (Amazona aestiva) and Orange-winged Amazon (Amazona amazonica) Parrots.

Safe and effective sedation protocols are important for chemical restraint of birds in clinical and diagnostic procedures, such as clinical evaluations, radiographic positioning, and blood collection. These protocols may reduce stress and ease the management of wild-caught birds, which are susceptible to injury or death when exposed to stressful situations. We compare the sedative effect of intranasal midazolam in wild-caught blue-fronted (Amazona aestiva) and orange-winged (Amazona amazonica) Amazon parrots. Ten adult parrots of each species (n = 20), of unknown sex, weighing 0.337 ± 0.04 (blue-fronted) and 0.390 ± 0.03 kg (orange-winged), kg were used. Midazolam (2 mg/kg) was administered intranasally and the total volume of the drug was divided equally between the 2 nostrils. Onset time and total sedation time were assessed. Satisfactory sedation for clinical evaluation was induced in all birds. Onset time and total sedation times were similar in both species: 5.36 ± 1.16 and 25.40 ± 5.72 minutes, respectively, for blue-fronted Amazons and 5.09 ± 0.89 and 27.10 ± 3.73 minutes, respectively, for orange-winged Amazons. A total of 15 animals showed absence of vocalization, with moderate muscle relaxation and wing movement upon handling, and 2 animals presented with lateral recumbence, with intense muscle relaxation and no wing movement, requiring no restraint. Three blue-fronted Amazons had no effective sedation. Intranasally administered midazolam at a dose of 2 mg/kg effectively promoted sedative effects with a short latency time and fast recovery in wild-caught parrots.

Histone methyltransferase SETD2 coordinates FACT recruitment with nucleosome dynamics during transcription.

Histone H3 of nucleosomes positioned on active genes is trimethylated at Lys36 (H3K36me3) by the SETD2 (also termed KMT3A/SET2 or HYPB) methyltransferase. Previous studies in yeast indicated that H3K36me3 prevents spurious intragenic transcription initiation through recruitment of a histone deacetylase complex, a mechanism that is not conserved in mammals. Here, we report that downregulation of SETD2 in human cells leads to intragenic transcription initiation in at least 11% of active genes. Reduction of SETD2 prevents normal loading of the FACT (FAcilitates Chromatin Transcription) complex subunits SPT16 and SSRP1, and decreases nucleosome occupancy in active genes. Moreover, co-immunoprecipitation experiments suggest that SPT16 is recruited to active chromatin templates, which contain H3K36me3-modified nucleosomes. Our results further show that within minutes after transcriptional activation, there is a SETD2-dependent reduction in gene body occupancy of histone H2B, but not of histone H3, suggesting that SETD2 coordinates FACT-mediated exchange of histone H2B during transcription-coupled nucleosome displacement. After inhibition of transcription, we observe a SETD2-dependent recruitment of FACT and increased histone H2B occupancy. These data suggest that SETD2 activity modulates FACT recruitment and nucleosome dynamics, thereby repressing cryptic transcription initiation.

Generation and characterization of induced pluripotent stem cell line (IBBISTi004-A) from an Angelman syndrome patient carrying a class II deletion of the maternal chromosome 15q11.2-q13.

Angelman Syndrome is a rare neurodevelopmental disorder caused by several (epi)genetic alterations. The patients present strong neurological impairment due to the absence of a functional maternal UBE3A gene in neurons. Here, we generated and characterized a new induced pluripotent stem cell (iPSC) line from a female child with Angelman syndrome harbouring a class II deletion. iPSCs were reprogrammed from fibroblasts using Sendai viruses. The new iPSCs express pluripotency markers, are capable of trilineage in vitro differentiation and have the expected imprinting status of Angelman syndrome. These iPSCs are a valuable tool to elucidate the pathophysiological mechanisms associated with this disease.

Birefringence Analyses Reveal Differences in Supramolecular Characteristics of Corneal Stromal Collagen Fibrils Between Falconiformes and Strigiformes.

This study aimed to assess the birefringent properties of corneal stromal collagen fibrils in birds of the orders Falconiformes (diurnal) and Strigiformes (predominantly nocturnal) to compare their supramolecular organizations. In total, 22 corneas of Falconiformes (Caracara plancus, n = 8; Rupornis magnirostris, n = 10; and Falco sparverius, n = 4) and 28 of Strigiformes (Tyto furcata, n = 16; Pseudoscops clamator, n = 6; and Athene cunicularia, n = 6) were processed histotechnically into 8-µm thick sections. Corneal optical retardation (OR) values related to the form and intrinsic fractions of the total birefringence of collagen fibrils were measured using a polarized light microscope equipped with phase compensators. In addition, the coherence coefficients that inform the local orientation of the fibrils were calculated through video image analysis. All assessments were conducted both in the anterior and posterior stroma of the cornea. Differences were significant when P < 0.05. The results showed supraorganizational differences between fibrils in the anterior stroma of Falconiformes and Strigiformes. The OR values were greater (P < 0.0001) for Falconiformes, indicating that the corneas of these birds contain more collagen fibrils or more aggregated collagen fibrils. In contrast, the coherence coefficients were higher (P = 0.016) for Strigiformes, indicating that the corneal collagen fibers in these birds are highly aligned and have few undulations. A multivariate data matrix constructed for Euclidean distance calculations showed that the dissimilarity between Falconiformes and Strigiformes corneas, in terms of the supraorganization of stromal collagen fibrils, was 4.56%. In conclusion, it is possible that the supraorganizational differences reported in this study may be sources of variation in the visual quality of Falconiformes and Strigiformes. This study provides the necessary evidence to encourage further research associating corneal optical performance to supramolecular characteristics of corneal stromal collagen.