Comparison of Intra-CL Injection and Peripheral Application of Prostaglandin F2α Analog on Luteal Blood Flow and Secretory Function of the Bovine Corpus Luteum

We investigated the effects of different doses of dinoprost injected directly into the bovine corpus luteum (CL) on (i) concentrations of progesterone (P4) and oxytocin (OT) in peripheral blood and (ii) mRNA levels of steroidogenic acute regulatory protein (STAR), cytochrome P450 family 11 subfamily A member 1 (P450scc), hydroxy-delta-5-steroid dehydrogenase, 3 β- and steroid delta-isomerase 1 (HSD3B), and receptor-interacting protein kinases 1 and 3 (RIPK1, RIPK3) in CL tissue. Moreover, we examined the effects of dinoprost, injected intra-CL or administered intramuscularly (IM), on CL secretory function and on indicators of CL vascular network status: luteal tissue area (LTA), CL blood flow (CLBF), and the CLBF:LTA ratio (Adj. CLBF), in cows at the early and mid-luteal phases. In the Experiment 1, cows (day 10 of the cycle) were allocated to (i) an intra-CL injection of saline (control; n = 3); (ii) an intra-CL injection of dinoprost (1.25 mg; 2.5 mg, or 5 mg; n = 3 for each dose); (iii) an IM administration of saline (control; n = 3); or (iv) an IM administration of dinoprost (25 mg; positive control; n = 3). Concentrations of OT and P4 were measured in plasma samples. The mRNA expression of steroidogenesis- or necroptosis-related factors was determined in CL tissue 4 h after treatments. In Experiment 2, cows on day 4 (n = 12) or day 10 (n = 12) were allocated to (i) an intra-CL injection of dinoprost (2.5 mg/0.5 ml; n = 6), or (ii) IM administration of dinoprost (25 mg/5 ml; n = 6). Concentrations of P4 were measured in plasma samples. Luteal tissue area, CLBF, and Adj. CLBF were assessed based on color Doppler ultrasonography. An intra-CL injection of dinoprost increased OT and decreased P4 levels in the jugular vein (JV) in a dose-dependent manner in cows at the mid-luteal phase. Increased CLBF and Adj. CLBF, accompanied by reduced P4 levels, were observed 2 h after intra-CL dinoprost injection in middle-stage CL. Decreased STAR and increased RIPK1 and RIPK3 mRNA levels confirmed that 2.5 mg dinoprost injected directly into CL is the minimum dose that induces luteolytic cascade. Injection of dinoprost directly into the CL (at a dosage lower than recommended for peripheral application) results in a pattern similar to IM dinoprost administration.

The Effectiveness of Compartmentalized Bone Graft Sponges Made Using Complementary Bone Graft Materials and Succinylated Chitosan Hydrogels

Bone defects can occur from many causes, including disease or trauma. Bone graft materials (BGMs) have been used to fill damaged areas for the reconstruction of diseased bone tissues since they are cost effective and readily available. However, BGMs quickly disperse around the tissue area, which ultimately leads to it migrating away from the defect after transplantation. We tested chitosan hydrogels as a useful carrier to hold BGMs in the transplantation area. In this study, we synthesized succinylated chitosan (SCS)-based hydrogels with a high decomposition rate and excellent biocompatibility. We confirmed that BGMs were well distributed inside the SCS hydrogel. The SCS-B hydrogel showed a decrease in mechanical properties, such as compressive strength and Young’s modulus, as the succinylation rate increased. SCS-B hydrogels also exhibited a high cell growth rate and bone differentiation rate. Moreover, the in vivo results showed that the SCS hydrogel resorbed into the surrounding tissues while maintaining the BGMs in the transplantation area for up to 6 weeks. These data support the idea that SCS hydrogel can be useful as a bioactive drug carrier for a broad range of biomedical applications.

Topographic Analysis of Low-Grade Myeloid Neoplasms By Multiparametric in Situ Imaging of Human Bone Marrow Core Biopsy Tissues

Diagnosis of low-grade myelodysplastic syndromes (LG-MDS) is one of the most challenging in hematopathology as it relies predominantly on morphologic assessment of dysplasia. Prior studies have demonstrated poor interobserver agreement among pathologists. Histomorphological evaluation of bone marrow core biopsy samples remains the gold standard for diagnostic workup of LG-MDS, including myelodysplastic syndromes (MDS) and other myeloid neoplasms. However, this approach may be subjective, and cannot quantitatively assess subtle differences in marrow topography and the cellular microenvironment. Multiparametric in situ imaging (MISI) through various techniques enables multiple biomarker detection in a single tissue. BostonGene has developed an AI-based image analysis platform to reveal spatial information and subtle histomorphologic features in an objective, quantitative fashion. Here, we demonstrate the potential for automated AI-based imaging analysis of MISI to assist in the differentiation of LG-MDS samples from normal marrow tissues (NBM). Decalcified human bone marrow core biopsy tissues from LG-MDS (n=6) and uninvolved staging marrows (NBM, n=4) were first prepared by immunofluorescence-based MISI via staining with DAPI and CD34, CD38, CD117, CD71, CD15, and CD61 antibodies. BostonGene analyzed the resulting images (fig.1) using a proprietary AI-based imaging platform to identify cells by segmentation performed with the pre-trained instance segmentation neural network. Cell types were identified with mean marker expression values using an accelerated version of BostonGene's phenograph clustering algorithm. Pathologists manually masked fat and bone trabeculae. Using a combination of cell size/shape parameters and antigen expression levels, the following unique cell types were identified: myeloblasts, proerythroblasts, erythroid normoblasts, maturing granulocytes, megakaryocytes, mast cells, plasma cells, and B-cell precursors (hematogones). Data revealed differences in the cellular content of NBM and LG-MDS samples, and separation of LG-MDS samples with the del(5q) subtype (n=2). While linear slender islet-like small clusters of erythroid normoblasts were detected in NBM, we observed a chaotic arrangement of them in LG-MDS samples. In LG-MDS samples, we found an increase in the total number of erythroid normoblasts from 17% to 31%, LG-MDS-del(5q) had 14%. The ratio of maturing granulocytes to erythroid normoblasts (M:E ratio) was significantly lower in LG-MDS (0.63) than in NBM (1.95). The M:E ratio generated by MISI strongly correlated to the M:E ratio produced by manual differential count of bone marrow aspirate samples (R=0.83, p < 0.003). Additionally, fewer hematogones were identified in LG-MDS marrows compared to NBM samples, as reported by others using orthogonal methods. Topographic analysis showed the fat to cellular tissue area ratio was higher in NBM (0.73) than LG-MDS (0.41), but the ratio of trabecular area to total tissue area was higher in LG-MDS (1.67) than NMB (0.74). Spatially, myeloblasts and megakaryocytes were found closer to trabeculae in NBM than LG-MDS;12 different cell communities were identified;2 of them (cluster 3 - erythroid normoblasts enriched, cluster 5 - erythroid normoblasts contacting proerythroblasts) were distributed statistically significantly differently in NBM and LG-MDS samples, indicating the use of MISI with AI-based imaging to distinguish LG-MDS from NBM. Patients of MDS-del(5q) subtype differ significantly from other MDS samples and are more similar to NBM. AI-based image analysis applied to MISI of bone marrow tissue revealed multiple cell types in single tissue sections, along with histologically subtle differences in topography between NBM and LG-MDS samples. These results highlight the importance of integrating in situ tissue analysis with techniques that examine single cell characteristics for a more comprehensive picture of the differences between normal tissue and tumor samples. Coupling sophisticated imaging analytics with this imaging method may provide a more powerful tool for novel biomarker discovery of prognostic and therapeutic significance in the management of MDS and other marrow-based disorders. Figure 1 Figure 1. Disclosures Svekolkin: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Varlamova: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Galkin: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Akaeva: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Smirnova: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Ovcharov: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Polyakova: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties. Tabakov: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company. Postovalova: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties: BostonGene. Koltakova: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company. Gunn: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company. Bagaev: BostonGene Corp.: Current Employment, Current holder of stock options in a privately-held company, Patents & Royalties: BostonGene.

An Insulin-like Growth Factor-1 Conjugated Bombyx mori Silk Fibroin Film for Diabetic Wound Healing: Fabrication, Physicochemical Property Characterization, and Dosage Optimization In Vitro and In Vivo

This study aimed to develop a silk fibroin (SF)-film for the treatment of chronic diabetic wounds. Silk fibroin was purified through a newly developed heating degumming (HD) process and casted on a hydrophobic surface to form SF-films. The process allowed the fabricated film to achieve a 42% increase in transparency and a 32% higher proliferation rate for BALB/3T3 fibroblasts compared to that obtained by conventional alkaline degumming treatment. Fourier transform infrared analysis demonstrated that secondary structure was retained in both HD- and alkaline degumming-derived SF preparations, although the crystallinity of beta-sheet in SF-film after the HD processing was slightly increased. This study also investigated whether conjugating insulin-like growth factor-1 (IGF-1) would promote diabetic wound healing and what the optimal dosage is. Using BALB/3T3 cells grown in hyperglycemic medium as a model, it was demonstrated that the optimal IGF-1 dosage to promote the cell growth was approximately 0.65 pmol. Further analysis of wound healing in a diabetic mouse model indicated that SF-film loaded with 3.25 pmol of IGF-1 showed significantly superior wound closure, a 13% increase at the 13th day after treatment relative to treatment with 65 pmol of free IGF-1. Improvement in diabetic wound healing was exerted synergistically by SF-film and IGF-1, as reflected by parameters including levels of re-epithelialization, epithelial tissue area, and angiogenesis. Finally, IGF-1 increased the epithelial tissue area and micro-vessel formation in a dose-dependent manner in a low dosage range (3.25 pmol) when loaded to SF-films. Together, these results strongly suggest that SF-film produced using HD and loaded with a low dosage of IGF-1 is a promising dressing for diabetic wound therapy.

Blocking cell fusion inhibits age-induced polyploidy and maintains epithelial organization in Drosophila

A characteristic of normal aging and age-related diseases is the remodeling of a tissue's cellular organization through polyploid cell growth. Polyploidy arises from an increase in nuclear ploidy or the number of nuclei per cell. However, it is not known whether age-induced polyploidy is an adaption to stressors or a precursor to degeneration. Here, we find that the adult fruit fly's abdominal epithelium becomes polyploid with age through generation of large multinucleated cells that make up more than 40% of the tissue area. The syncytia arise by cell fusion, not endomitosis. Epithelial multinucleation is also a characteristic of macular degeneration, including Ctnna1tvrm5, a mouse model for pattern dystrophy. Similarly, we find that the knockdown of alpha-catenin enhances multinucleation in the fly epithelium. We further show that age-induced polyploidy can be suppressed by inhibiting cell fusion revealing a means to maintain tissue organization in older animals.

Radial variability of fibrovascular bundle properties of salacca (Salacca zalacca) fronds cultivated on Turi Agrotourism in Yogyakarta, Indonesia

Hakim L, Widyorini R, Nugroho WD, Prayitno TA. 2021. Radial variability of fibrovascular bundle properties of salacca (Salacca zalacca) fronds cultivated on Turi Agrotourism in Yogyakarta, Indonesia. Biodiversitas 22: 3594-3603. Fibrovascular bundles have properties variability not only based on species and varieties but also parts of species. This study, therefore, aims to characterize the FVB fundamental properties (anatomical, chemical, physical and mechanical) of Salacca zalacca (Gaertn.) Voss fronds, based on radial direction. The salacca fronds were divided into three parts, outer, middle as well as inner positions. Then the FVB's anatomical and physical properties were observed by light microscope and gravimetry analysis, respectively. Meanwhile, the variability of chemical and mechanical properties was investigated based on the ASTM standard. According to the results, the outer position has a higher variability of diameter, density, cellulose, lignin, and mechanical properties than the inner position, but has a lower hemicellulose value than the middle and inner position. Furthermore, the relationships between the anatomical, physical, chemical, and mechanical properties were discovered to form a pattern where increasing the mechanical properties is influenced by density and ratio vascular tissue area to total transverse area. Based on the results, the fibrovascular bundle of S. zalacca frond was concluded to possess anatomical, physical, chemical, and mechanical properties variability on the radial direction. There was a correlation between anatomical properties and mechanical properties.

The correlation between type 2 diabetes and fat fraction in liver and pancreas: a study using MR Dixon technique

Background The increased obesity results in ectopic fat deposits in liver and pancreas.Ectopic fat deposits affect insulin resistance and blood sugar content with Type 2 Diabetes.To assess the relationship between obesity and ectopic fat deposits and diabetes,this study used MR Dixon method for the quantification of liver and pancreas fat fraction (FF) in T2DM patients and healthy controls. Methods The FF of liver and pancreas, the maximum diameters of the pancreas, SAT, VAT and TAT were measured for 167 subjects using the MR Dixon data.Four groups were established on the basis of BMI value.For statistics, intra-and inter-group comparison was done by employing Independent Sample t Test. Results ① The average fat content in liver and pancreas, the fat content in pancreas body and tail, and VAT in Group 1 were higher than those in Group 3 (P༜0.05). ② The average fat content in liver and pancreas, the adipose fraction of the pancreas head, and VAT in the Group 2 were higher than those in Group 4 (P༜0.05). ③ The average fat content of liver and pancreas, the adipose content of the body and the tail of the pancreas, the abdominal subcutaneous adipose area (SAT), the visceral adipose tissue area (VAT), and the total abdominal adipose tissue area (TAT) in Group 2 were higher than those in the Group 1 (P༜0.05). ④ The FF of pancreas tail, SAT and TAT in Group 4 were higher than those in Group 3 (P༜0.05). Conclusion The tissue FF,which has a close relationship with T2DM,can be assessed by MR Dixon technique.The results showed that all T2DM patients should pay attention to tissue fat content regardless of BMI values.

Ultrasound Characteristics of the Cavitary Corpus Luteum after Oestrus Synchronization in Heifers in Relation to the Results of Embryo Transfer

The aim of the study was to conduct an ultrasound analysis of quantitative parameters of the corpus luteum (CL) in recipient heifers on days 6–8 after oestrus, and to compare reproduction potential of both types of CL in those females. Analyses were performed on 300 heifers, synchronized with two injections of cloprostenol. Clinical and ultrasound examinations of ovaries were performed and measurements of the CL were recorded. The blood samples were taken to determine progesterone level. Pregnancy examination was conducted after 6–8 weeks from the ET. Cavitary CL was found in 32.7% heifers In 48.0% of the cavitary CL, its luteal tissue area was reduced by 14.3% compared to the compact CL, while 16.3% of the CL had luteal tissue reduced by more than 33.8%. Progesterone level in blood serum was higher in heifers with the cavitary CL (p < 0.001). Pregnancy rate was higher for females with a cavitary CL (52%) than those with compact ones (33%, p < 0.05). The ultrasound assessment of luteal tissue should be included in the evaluation of the functional status of the CL in ET-recipient heifers. The cavitary CL presence may indicate a higher potential of the recipient in maintaining the pregnancy.

Association of BMI, body composition and outcomes in Chinese patients with metastatic renal cell carcinoma treated with immunotherapy: A retrospective, multicohort analysis.

e16563 Association of BMI, body composition and outcomes in Chinese patients with metastatic renal cell carcinoma treated with immunotherapy: a retrospective, multicohort analysis. Background: Obesity (body-mass index [BMI] ≥30 kg/m) is associated with a higher risk of developing clear cell renal cell carcinoma (RCC) but, paradoxically, obesity is also associated with better outcome in renal cell carcinoma (RCC) patients treated with immunotherapy. Whether BMI associate outcomes in Chinese RCC patients treated with immunotherapy considering Asian population has a lower BMI? In this study, we aimed to investigate the relationship between BMI, body composition and outcomes of Chinese patients with RCC treated with immunotherapy. Methods: We evaluated clinical data of metastatic RCC patients treated with immunotherapy from five major centers in China. Despite using the Chinese standard, only 6 people met the obese standard (BMI ≥28 kg/m, as per WHO's BMI categories), so we divided the patients into high BMI gourps (BMI ≥24.0 kg/m), and normal weight groups (BMI 18.5-23·9 kg/m). We assessed overall survival, defined as the time from treatment initiation to the date of any-cause death or of censoring on the day of the last follow-up, in high BMI patients and in patients with a normal weight. We also investigative the relationship between body composition (skeletal muscle index (SMI = skeletal muscle area/height2), skeletal muscle density (SMD), and subcutaneous adipose distribution index (SADI = subcutaneous adipose tissue area / subcutaneous adipose tissue area + visceral adipose tissue area) of each patient at the third lumbar vertebrae) and overall survival using CT or MRI scan at the treatment initiation. Results: Out of a total of 222 patients, 203 had evaluable CT or MRI radiographs, pretreatment BMI measurements, and overall survival data for analyses, and we excluded 4 (1.7%) underwetight patients, leaving a final cohort of 199 patients from this study for our analyses. There were 79 (39.7%) high BMI patients and 120 (65.3%) normal weight patients. There was no difference in overall survival between the high BMI and normal weight groups (HR:1.36, CI 0.83-2.21). In addition, SMI and SMD were not associated with outcome of patients. Interestingly, although there was no statistical difference, subcutaneous fat was associated with better outcomes, while visceral fat was associated with poor outcomes. Thus, the combination of the two indicators, SADI, showed a significant prognostic correlation after adjustment for International Metastatic RCC Database risk score (aHR:0.314, CI 0.19-0.51). Conclusions: BMI is not a good predictor of the outcome of immunotherapy in Chinese patients with RCC. Evaluation of body composition may be a better tool for predicting prognosis in RCC patients treated with immunotherapy.