Neuron Navigator 3 (NAV3) is Required for Heart Development in Zebrafish

As a tightly controlled biological process, cardiogenesis requires the specification and migration of a suite of cell types to form a particular three-dimensional configuration of the heart. Many genetic factors are involved in the formation and maturation of the heart, and any genetic mutations may result in severe cardiac failures. The neuron navigator (NAV) family consists of three vertebrate homologs (NAV1, NAV2, and NAV3) of the neural guidance molecule Uncoordinated-53 (UNC-53) in Caenorhabditis elegans. Although they are recognized as neural regulators, their expressions are also detected in many organs, including the heart, kidney, and liver. However, the functions of NAVs, regardless of neural guidance, remain largely unexplored. In our study, we found that nav3 gene was expressed in the cardiac region of zebrafish embryos from 24 to 48 hours post-fertilization (hpf) by means of in situ hybridization (ISH) assay. A CRISPR/Cas9-based genome editing method was utilized to delete the nav3 gene in zebrafish and loss-of-function of Nav3 resulted in a severe deficiency in its cardiac morphology and structure. The similar phenotypic defects of the knockout mutants could recur by nav3 morpholino injection and be rescued by nav3 mRNA injection. Dual-color fluorescence imaging of ventricle and atrium markers further confirmed the disruption of the heart development in nav3-deleted mutants. Although the heart rate was not affected by the deletion of nav3, the heartbeat intensity was decreased in the mutants. All these findings indicate that Nav3 was required for cardiogenesis in developing zebrafish embryos.

373 Utility of the early phase of DPD scintigraphy in the diagnosis of correlated TT-R cardiac amyloidosis

Aims The importance of cardiac scan with phosphonate-based radiotracers in the diagnosis of cardiac amyloidosis is now well established. Standard imaging is performed 3 h after tracer injection with a planar view on the cardiac region. This study sought to evaluate the predictive role of early-phase myocardial uptake (10 min after injection) of 99mTc-3,3-diphosphono-1,2-propanodicarboxylic acid (DPD) compared compared with standard late acquisition, in patients with suspected hereditary transthyretin-related cardiac amyloidosis (TTR-CA). Methods and results Fifty five patients with suspected of TTR-CA with typical aspects of the relative apical sparing at two-dimensional speckle-tracking echocardiography, reported as a specific pattern for cardiac amyloidosis, were enrolled after having signed informed written consent. They have been subjected to a 99mTc-DPD cardiac scintigraphy with planar acquisition at 10 min and 3 h after tracer injection (13 Mbq/Kg). Patients with cardiac uptake on the planar images concluded the examination with a SPECT-CT (cardiac protocol) to assess the affected myocardial segments. On planar images the heart-to-mediastinum-ratio was measured. Subsequently, the diagnosis of amyloidosis has to be confirmed with morphologic examinations such as biopsy and genetic tests. Of the enrolled patients with clinical and echocardiographic aspect of TTR-CA, 22 were positive for cardiac amyloidosis. All of them showed tracer uptake in both early and late images. In patients with positive results, the early-phase showed a Heart-to-mediastinum-ratio >1.2. SPECT/CT showed involvement of almost two myocardial segments: in all patients the ventricular septum showed significant tracer uptake. Conclusions Our small group of patients showed that 99mTc-DPD myocardial uptake intensity on early-phase scintigraphy can be used to anticipate the results of late images in diagnosis of TTR cardiac amyloidosis.

ASB2 is a novel E3 ligase of SMAD9 required for cardiogenesis

Cardiogenesis requires the orchestrated spatiotemporal tuning of BMP signalling upon the balance between induction and counter-acting suppression of the differentiation of the cardiac tissue. SMADs are key intracellular transducers and the selective degradation of SMADs by the ubiquitin–proteasome system is pivotal in the spatiotemporal tuning of BMP signalling. However, among three SMADs for BMP signalling, SMAD1/5/9, only the specific E3 ligase of SMAD9 remains poorly investigated. Here, we report for the first time that SMAD9, but not the other SMADs, is ubiquitylated by the E3 ligase ASB2 and targeted for proteasomal degradation. ASB2, as well as Smad9, is conserved among vertebrates. ASB2 expression was specific to the cardiac region from the very early stage of cardiac differentiation in embryogenesis of mouse. Knockdown of Asb2 in zebrafish resulted in a thinned ventricular wall and dilated ventricle, which were rescued by simultaneous knockdown of Smad9. Abundant Smad9 protein leads to dysregulated cardiac differentiation through a mechanism involving Tbx2, and the BMP signal conducted by Smad9 was downregulated under quantitative suppression of Smad9 by Asb2. Our findings demonstrate that ASB2 is the E3 ligase of SMAD9 and plays a pivotal role in cardiogenesis through regulating BMP signalling.

Convolutional neural network-based automatic heart segmentation and quantitation in 123I-metaiodobenzylguanidine SPECT imaging

Background Since three-dimensional segmentation of cardiac region in 123I-metaiodobenzylguanidine (MIBG) study has not been established, this study aimed to achieve organ segmentation using a convolutional neural network (CNN) with 123I-MIBG single photon emission computed tomography (SPECT) imaging, to calculate heart counts and washout rates (WR) automatically and to compare with conventional quantitation based on planar imaging. Methods We assessed 48 patients (aged 68.4 ± 11.7 years) with heart and neurological diseases, including chronic heart failure, dementia with Lewy bodies, and Parkinson's disease. All patients were assessed by early and late 123I-MIBG planar and SPECT imaging. The CNN was initially trained to individually segment the lungs and liver on early and late SPECT images. The segmentation masks were aligned, and then, the CNN was trained to directly segment the heart, and all models were evaluated using fourfold cross-validation. The CNN-based average heart counts and WR were calculated and compared with those determined using planar parameters. The CNN-based SPECT and conventional planar heart counts were corrected by physical time decay, injected dose of 123I-MIBG, and body weight. We also divided WR into normal and abnormal groups from linear regression lines determined by the relationship between planar WR and CNN-based WR and then analyzed agreement between them. Results The CNN segmented the cardiac region in patients with normal and reduced uptake. The CNN-based SPECT heart counts significantly correlated with conventional planar heart counts with and without background correction and a planar heart-to-mediastinum ratio (R2 = 0.862, 0.827, and 0.729, p < 0.0001, respectively). The CNN-based and planar WRs also correlated with and without background correction and WR based on heart-to-mediastinum ratios of R2 = 0.584, 0.568 and 0.507, respectively (p < 0.0001). Contingency table findings of high and low WR (cutoffs: 34% and 30% for planar and SPECT studies, respectively) showed 87.2% agreement between CNN-based and planar methods. Conclusions The CNN could create segmentation from SPECT images, and average heart counts and WR were reliably calculated three-dimensionally, which might be a novel approach to quantifying SPECT images of innervation.

VAJRA KANJIKA – ITS BENEFITS ON SUTIKA

A woman immediately after giving birth to a child is called Sutika. Various Acharya's have mentioned Sutika Kala as a very important period because the mother is prone to many complications following the delivery, where a lot of Rakta and Kleda are lost in the process of delivery leaving her vulnerable for various disorders. As we know Vata fills the Rikta Sthana and causes various disorders, in a similar way Sutika is very much prone to Vata disor- ders. Mainly we come across disorders like inadequacy of breast milk and Sutika Makkala (pain). Sthanya is very important in the growth and development of the newborn. It enhances the immunity of the child. Breast milk is babies specific hence one should adopt measures to improve lactation instead of depending on supplementary feeds. Another important condition is Sutika Makkala which is caused due to vitiation of Vata, and shows symptoms like severe discomfort, pain below the umbilical and bladder region, abdomen and cardiac region. At times the pain is so severe that a prickly sensation with a possibility of tearing of bowel or its perforation. These are associated with flatulence and retention of urine. In these above conditions, Vajra Kanjika is of utmost benefit. We get the reference of Vajrakanjika in Yogaratnakara in Ksheera Dosha Chikitsa Adhyaya. Vajra Kanjika contains the drugs like Pip- pali, Pippali mula, Chavya, Shunti, Yawanika, both Jeeraka’s, both Haridra’s, Vida and Souvarchala Lavana. It is indicated in Amavata, it is Vrishya, suppresses Kapha and Vata, increases appetite, cures Makkala Shula and in- creases the quantity of milk.In this article, an attempt is made to understand the probable mode of action of Vajra Kanjika in Sutika, Sutika Makkala and Sthanya kshaya. Keywords: Vajra Kanjika, Sutika, Sutika Roga, Makkala, Sthanya Dosha.

ULTRASTRUCTURAL STUDY OF THE ESOPHAGUS AND STOMACH OF Arapaima gigas (Schinz, 1822), JUVENILE PAICHE, CREATED EXCAVATED TANK

Paiche ( Arapaima gigas) belongs to the Kingdom Animalia , Phylum Chordata , Class Actinopterygii , Order Osteoglossiformes, Family Arapaimidae, Genus Arapaima, and its origin may date to the Jurassic period. The species has natural habitat in the Amazonian rivers, found mainly in marginal lakes, being considered an important fishing resource, with high market value and high demand for meat and leather in both Brazilian and international trade. This study aims to describe the morphology of the esophagus and stomach by light microscopy and scanning electronics microscopy. The esophagus was presented as muscular, short, tubular and fan-shaped in the cranial portion, also presenting deep longitudinal folds, and the entire mucosa is covered by mucus secretory cells with distinct morphological characteristics. Pirarurcu’s stomach has a J-shape divided into three regions: cardiac with a lighter aspect, fundus portion with few folds in the mucosa, and pyloric with deeper folds, also presenting gastroliths in fundus and pyloric portions. Both microscopy studies highlighted three glandular regions, composed by mucoid columnar epithelial cells, gastric crypts with different shapes and sizes depending on each portion, in which the different shapes of the mucosal folds in each region of the stomach were evident, and digitiform microsaliences were found in the cardiac region, and micro-orifices and desmosome in the fundus region. Also, fundus and pyloric portions produce more mucus than the cardiac. Then morphology found was consistent with the eating habits and management of distinct characteristics of the digestive tract.

Assessing Doxorubicin-Induced Cardiomyopathy by 99mTc-3PRGD2 Scintigraphy Targeting Integrin αvβ3 in a Rat Model

The present study evaluated interstitial alterations in doxorubicin-induced cardiomyopathy using a radiolabeled RGD peptide 99mTc-3PRGD2 specific for integrin αvβ3 that targets myofibroblasts.Cardiomyopathy was induced in 20 Sprague-Dawley rats by intraperitoneal doxorubicin injections (2.5 mg/kg/week) for up to six weeks. 99mTc-3PRGD2 scintigraphy was performed in control rats (n = 6) at baseline and three, six, and nine weeks after first doxorubicin administration (n = 6, 6, and 5 for each time point). For another three rats of 6-week modeling, cold c(RGDyK) was co-injected with 99mTc-3PRGD2 to evaluate specific radiotracer binding. Semi-quantitative parameters were acquired to compare radiotracer uptake among all groups. The biodistribution of 99mTc-3PRGD2 was evaluated by a γ-counter after scintigraphy. Haematoxylin and eosin, and Masson’s staining were used to evaluate myocardial injury and fibrosis, while western blotting and immunofluorescence co-localization were used to analyze integrin αvβ3 expression in the myocardium.The 99mTc-3PRGD2 half-life in the cardiac region (Heartt 1/2) of the 9-week model and heart radioactivity percentage (%Heart20 min, %Heart40 min and %Heart60 min) of the 6 and 9-week models were significantly increased compared to the control. Heart-to-background ratio (HBR20 min, HBR40 min and HBR60 min) increase began in the third week, continued until the sixth week, and was reversed in the ninth week, which paralleled the changing trend of cardiac integrin αvβ3 expression. The myocardial biodistribution of 99mTc-3PRGD2 was significantly correlated with integrin β3 expression.The 99mTc-3PRGD2 scintigraphy allows for non-invasive visualization of interstitial alterations during doxorubicin-induced cardiomyopathy.

Histochemical Assessment of Mucin-Secreting Cells from the Gastric Mucosa of Guinea Pigs

Stomach fragments from 3 guinea pigs were collected from the three regions: cardiac, fundic and pyloric, for histochemical investigations. The anatomical segments were processed by the classical method of inclusion in paraffin and the histological sections were stained with PAS reaction for highlighting the neutral mucins and the Alcian blue method for acid mucins. All the surface cells of the gastric mucosa and in the crypts were positive on the PAS reaction and negative in the case of the alcian blue reaction. This demonstrates that cells on the surface and in the crypts synthetize neutral mucins. In the case of the cardiac region glands, only a small number of cells were positive on the two histochemical reactions, which shows that the cardiac glands in guinea pig synthesize a very small amount of neutral and acidic mucins. There are no positive cells in neither reaction used in the fundic glands case, which shows that these glands do not synthesize mucins, neither neutral nor acidic content. The glands located in the pyloric region have cells in the deep half of the wall which were positive on both histochemical reactions, which shows that they synthesize both neutral mucins and acidic mucins.

Immediate and Long-Term Results of the Spleen-Preserved Operations in the Surgical Treatment of Gastric Cancer

ntroduction. Currently, total gastrectomy with D2 lymphadenectomy is the standard surgical treatment for proximal gastric cancer at the resectable stages (I-III). The issue of advisability of splenectomy as a component of lymphadenectomy remains a controversial one, especially when the tumor is localized in the region of the body or cardiac region of the stomach.The aim of the study was to compare immediate and long-term outcomes, including the quality of life, between spleen preserving and spleen removing surgeries.Methods. The study included 363 patients with gastric cancer II-III stages, localized in the upper and/or the middle third of the stomach, who underwent surgery at the Voronezh Regional Clinical Oncology Hospital and the Voronezh Clinical Hospital of the Russian Railway-Medicine in 2015-2017. All patients were conditionally divided into 2 groups for comparative retrospective analysis. All patients of the first (experimental or spleen-preserved) group (144 patients) were performed R0 total gastrectomy with D2 lymphadenectomy, including splenic hilar nodes (№ 10,11) removal without splenectomy. Patients of the second (control or splenectomy) group (219 patients) were performed R0 total gastrectomy with D2 lymphadenectomy and prophylactic splenectomy (for splenic hilar nodes removal).Results. The average duration of the operation and the volume of blood loss did not differ in both groups. The incidence of early postoperative surgical complications was lower in the spleen-preserved group. Splenectomy was associated with more severe complications of class 4 and 5 according to the Clavien-Dindo classification. Conclusion. Parameters of the 1- and 3-year overall survival rate did not differ in both groups. The results of the GSRS questionnaire were similar in both groups, excluding reflux-esophageal symptoms scale. The reflux scale demonstrated a statistically and clinically significant advantage of spleen preservation.