Systemic amyloid A amyloidosis of the bladder after transurethral resection of urothelial carcinoma.

Introduction: Amyloid A amyloidosis of the bladder is not a major disease. We report a patient with systemic amyloid A amyloidosis of the bladder after transurethral resection of urothelial carcinoma. Case presentation: An 87-year-old Japanese man had bladder carcinoma. He was followed up regularly with cystoscopy. Cystoscopy revealed multiple polypoid tumors 6 months after the first transurethral resection of urothelial carcinoma. Pathologic specimens contained the amyloid A component. He had hypertrophic cardiomyopathy, valvular disorders, and arrhythmias. His cardiac disease may have resulted from amyloid A amyloidosis. We speculated the patient had systemic amyloid A amyloidosis of the heart and bladder. Conclusion: We determined the type of amyloidosis via a biopsy of the bladder tumors. Our patient had cardiac disease. Therefore, systemic amyloid A amyloidosis could have caused his cardiac disease. The pathologic findings of bladder tumors can contribute to detecting systemic amyloid A amyloidosis.

Rare Coexistence of Aldosterone-producing Adrenocortical Adenoma Confirmed by an Immunohistochemical Analysis of Steroidogenic Enzymes with Adrenal Ectopic Thyroid Tissue: A Case Report and Literature Review.

A 56-year-old man presented with a history of hypertension; clinically, the patient had primary aldosteronism (PA) and a 4-cm left adrenal tumor. The left adrenal glands, resected by adrenalectomy, also contained ectopic thyroid tissue (ETT). An immunohistochemical analysis of steroid-converting enzymes revealed an aldosterone-producing adenoma (APA). Among 19 previously reported cases of adrenal ETT, 4 had adrenal hormonal abnormalities, all of which were PA. This is the first case of adrenal ETT coexisting with APA, confirmed by steroid-converting enzyme expression. Further analyses using cumulative case data are required to clarify the correlation between adrenal ETT and APA.

Clonal Origin and Lineage Ambiguity in Mixed Neuroendocrine Carcinoma of the Uterine Cervix.

Small-cell neuroendocrine carcinoma (SCNEC) of the cervix is a rare disease characterized by a high incidence of mixed tumors with other types of cancer. The mechanism underlying this mixed phenotype is not well understood. This study established a panel of organoid lines from patients with SCNEC of the cervix and ultimately focused on one line, which retained a mixed tumor phenotype, both in vitro and in vivo. Histologically, both organoids and xenograft tumors showed distinct differentiation into either SCNEC or adenocarcinoma in some regions and ambiguous differentiation in others. Tracking single cells indicated the existence of cells with bipotential differentiation toward SCNEC and adenocarcinomas. Single-cell transcriptional analysis identified three distinct clusters: SCNEC-like, adenocarcinoma-like, and a cluster lacking specific differentiation markers. The expression of neuroendocrine markers was enriched in the SCNEC-like cluster but not exclusively. Human papillomavirus 18 E6 was enriched in the SCNEC-like cluster, which showed higher proliferation and lower levels of the p53 pathway. After treatment with anticancer drugs, the expression of adenocarcinoma markers increased, whereas that of SCNEC decreased. Using a reporter system for keratin 19 expression, changes in the differentiation of each cell were shown to be associated with the shift in differentiation induced by drug treatment. These data suggest that mixed SCNEC/cervical tumors have a clonal origin and are characterized by an ambiguous and flexible differentiation state.

Positive Regulation of S-Adenosylmethionine on Chondrocytic Differentiation via Stimulation of Polyamine Production and the Gene Expression of Chondrogenic Differentiation Factors.

S-adenosylmethionine (SAM) is considered to be a useful therapeutic agent for degenerative cartilage diseases, although its mechanism is not clear. We previously found that polyamines stimulate the expression of differentiated phenotype of chondrocytes. We also found that the cellular communication network factor 2 (CCN2) played a huge role in the proliferation and differentiation of chondrocytes. Therefore, we hypothesized that polyamines and CCN2 could be involved in the chondroprotective action of SAM. In this study, we initially found that exogenous SAM enhanced proteoglycan production but not cell proliferation in human chondrocyte-like cell line-2/8 (HCS-2/8) cells. Moreover, SAM enhanced gene expression of cartilage-specific matrix (aggrecan and type II collagen), Sry-Box transcription factor 9 (SOX9), CCN2, and chondroitin sulfate biosynthetic enzymes. The blockade of the methionine adenosyltransferase 2A (MAT2A) enzyme catalyzing intracellular SAM biosynthesis restrained the effect of SAM on chondrocytes. The polyamine level in chondrocytes was higher in SAM-treated culture than control culture. Additionally, Alcian blue staining and RT-qPCR indicated that the effects of SAM on the production and gene expression of aggrecan were reduced by the inhibition of polyamine synthesis. These results suggest that the stimulation of polyamine synthesis and gene expression of chondrogenic differentiation factors, such as CCN2, account for the mechanism underlying the action of SAM on chondrocytes.

Localized Amyloid A amyloidsis of ureter: A case report.

Amyloidosis of the ureter is a rare disease, distinguishing it from a neoplasm is difficult. A 64-year-old Japanese woman suffered from macrohematuria and left side hydronephrosis with ultrasound in 2014. Retrograde pyelography revealed no ureter tumor at that time. The patient had macrohematuria, left side hydronephrosis, and ureteral stenosis in the left ureter on retrograde pyelography. She was suspected of having a ureter tumor when she was 71 years old in 2021. The patient underwent ureteroscopy with biopsy. Pathological specimens contained the amyloid A component, based on immunohistochemistry staining. We diagnosed the patient with amyloid A amyloidosis of the ureter.

Dual roles of cellular communication network factor 6 (CCN6) in the invasion and metastasis of oral cancer cells to bone via binding to BMP2 and RANKL.

The acquisition of motility via epithelial-mesenchymal transition (EMT) and osteoclast induction are essential for the invasion and metastasis of oral squamous cell carcinoma (OSCC) to bone. However, the molecule suppressing both EMT and osteoclastogenesis is still unknown. In this study, we found that cellular communication network factor 6 (CCN6) was less produced in a human OSCC cell line, HSC-3 with mesenchymal phenotype, than in HSC-2 cells without it. Notably, CCN6 interacted with bone morphogenetic protein 2 (BMP2) and suppressed the cell migration of HSC-3 cells stimulated by BMP2. Moreover, knockdown of CCN6 in HSC-2 cells led to the promotion of EMT and enhanced the effect of transforming growth factor-ß (TGF-ß) on the promotion of EMT. Furthermore, CCN6 combined with BMP2 suppressed EMT. These results suggest that CCN6 strongly suppresses EMT in cooperation with BMP2 and TGF-ß. Interestingly, CCN6 combined with BMP2 increased the gene expression of receptor activator of nuclear factor-κB ligand (RANKL) in HSC-2 and HSC-3 cells. Additionally, CCN6 interacted with RANKL, and CCN6 combined with RANKL suppressed RANKL-induced osteoclast formation. In metastatic lesions, increasing BMP2 due to the bone destruction led to interference with binding of CCN6 to RANKL, which results in the promotion of bone metastasis of OSCC cells due to continuous osteoclastogenesis. These findings suggest that CCN6 plays dual roles in the suppression of EMT and in the promotion of bone destruction of OSCC in primary and metastatic lesions, respectively, through cooperation with BMP2 and interference with RANKL.

Omega-3 meat enrichment and L-FABP, PPARA, and LPL genes expression are modified by the level and period of tuna oil supplementation in slow-growing chickens.

This study proposes a strategy to manipulate the fatty acid (FA) content in slow-growing Korat chicken (KRC) meat using tuna oil (TO). To determine the optimal level and feeding period of TO supplementation, we conducted a study investigating the effects of dietary TO levels and feeding periods on meat quality, omega-3 polyunsaturated fatty acid (n-3 PUFA) composition, and gene expression related to FA metabolism in KRC breast meat. At 3 wk of age, 700 mixed-sex KRC were assigned to seven augmented factorial treatments with a completely randomized design, each consisting of four replicate pens containing 25 chickens per pen. The control group received a corn-soybean-based diet with 4.5% rice bran oil (RBO), while varying amounts of TO (1.5%, 3.0%, or 4.5%) replaced a portion of the RBO content in the experimental diets. The chickens were fed these diets for 3 and 6 wk, respectively, before being slaughtered at 9 wk. Our results indicated no significant interactions between TO levels and feeding periods on the growth performance or meat quality of KRC (P > 0.05). However, the liver fatty acid-binding protein gene (L-FABP, also known as FABP1), responsible for FA transport and accumulation, showed significantly higher expression in the chickens supplemented with 4.5% TO (P < 0.05). The chickens supplemented with 4.5% TO for a longer period (3 to 9 wk of age) exhibited the lowest levels of n-6 PUFA and n-6 to n-3 ratio, along with the highest levels of eicosapentaenoic acid, docosahexaenoic acid, and n-3 PUFA in the breast meat (P < 0.05). However, even a short period of supplementation with 4.5% TO (6 to 9 wk of age) was adequate to enrich slow-growing chicken meat with high levels of n-3 PUFA, as recommended previously. Our findings indicated that even a short period of tuna oil supplementation could lead to desirable levels of omega-3 enrichment in slow-growing chicken meat. This finding has practical implications for the poultry industry, providing insights into optimal supplementation strategies for achieving desired FA profiles without adversely affecting growth performance or meat quality.

This study investigated the effect of different levels and feeding periods of tuna oil (TO), a source of omega-3 polyunsaturated fatty acids (n-3 PUFA), was used to modify the fatty acid (FA) profile in slow-growing Korat chicken (KRC) meat. The interaction between TO supplementation levels and feeding periods did not influence growth performance or meat quality in KRC. However, higher level of TO supplementation led to increased expression of the liver fatty acid-binding protein gene, which is involved in FA transport and accumulation. The highest levels of eicosapentaenoic acid, docosahexaenoic acid, and n-3 PUFA were detected in the chickens that were fed 4.5% TO supplementation for a long period (3 to 9 wk of age). These chickens also had the lowest amounts of omega-6 polyunsaturated fatty acids (n-6 PUFA) and n-6 to n-3 ratio. Interestingly, even a short period of 4.5% TO supplementation (6 to 9 wk of age) in slow-growing chickens was sufficient to enrich the KRC meat with n-3 PUFA. These findings highlight the potential for improving the nutritional profile of chicken meat by regulating TO supplementation in the diet.

Transcriptome analysis of the uterovaginal junction containing sperm storage tubules in heat-stressed breeder hens.

Sperm storage tubules (SSTs) in the uterovaginal junction (UVJ) of the oviduct are major sites of sperm storage after artificial insemination or mating. Female birds may regulate sperm motility in the UVJ. Heat stress can decrease the reproductive ability of broiler breeder hens. However, its effects on UVJ remain unclear. Changes in gene expression aid in understanding heat stress-affected molecular mechanisms. Herein, we wanted to conduct a comparative transcriptomic analysis to identify the differentially expressed genes (DEGs) in the UVJ of breeder hens under thermoneutral (23°C) and heat stress (36°C for 6 h) conditions. The results indicated that cloacal temperatures and respiratory rates were significantly increased in heat-stressed breeder hens (P < 0.05). Total RNA was extracted from the hen UVJ tissues containing SSTs after heat exposure. Transcriptome analysis identified 561 DEGs, including 181 upregulated DEGs containing heat shock protein (HSP) transcripts and 380 downregulated DEGs containing immune-related genes, such as interleukin 4-induced 1, radical S-adenosyl methionine domain containing 2, and 2'-5'-oligoadenylate synthetase like, in heat-stressed hens. Gene Ontology analysis revealed the significantly enriched terms involving HSPs. Kyoto Encyclopedia of Genes and Genomes analysis identified 9 significant pathways, including the protein processing in endoplasmic reticulum (11 genes including HSPs), neuroactive ligand-receptor interaction (13 genes including luteinizing hormone/choriogonadotropin receptor), biosynthesis of amino acids (4 genes including tyrosine aminotransferase), ferroptosis (3 genes including heme oxygenase 1), and nitrogen metabolism (carbonic anhydrase [CA]-12 and CA6) pathways. Protein-protein interaction network analysis of DEGs revealed 2 large networks, one containing upregulated HSPs and the other containing downregulated interferon-stimulating genes. Overall, heat stress inhibits innate immunity in the UVJ tissues of broiler chickens, and heat-stressed chickens protect their cells by increasing the expression levels of HSPs. The identified genes are potential candidates for further exploration of the UVJ in heat-stressed hens. The identified molecular pathways and networks increase our understanding of the sperm storage reservoirs (UVJ containing SSTs) within the reproductive tract and may be used to prevent heat stress-induced fertility loss in breeder hens.

Protocols for Screening Peptides Binding to CCN Family Proteins and Their Extended Utility.

The function of CCN family proteins is determined by their interactions with multiple cofactors that are present in the microenvironment. Therefore, determining these cofactors is critically important in understanding the molecular function of CCN family members. For this objective, a bacteriophage random peptide display library is a suitable tool. In this library, each filamentous bacteriophage is designed to display an oligopeptide of 7-20 random amino acid residues on its surface. Bacteriophage clones that possess peptides that bind to a CCN family protein are selected through several cycles of a process called biopanning or affinity selection. By determining the nucleotide sequence of the DNA that encodes the displayed peptide, the oligopeptides that specifically bind to the CCN family member can be specified. The obtained peptide sequences can be utilized to design peptide aptamers for CCN family proteins, or as a key sequence to determine new CCN family cofactor candidates in silico. Instead of a random peptide cDNA library, an antibody cDNA library from naïve lymphocytes or from B cells immunized by a CCN family protein can be used in order to obtain a highly specific CCN family detection or functional modulation tool.

Cysteinyl leukotriene receptor 1 is dispensable for osteoclast differentiation and bone resorption.

Cysteinyl leukotriene receptor 1 (CysLTR1) is a G protein-coupled receptor for the inflammatory lipid mediators cysteinyl leukotrienes, which are involved in smooth muscle constriction, vascular permeability, and macrophage chemokine release. The Cysltr1 gene encoding CysLTR1 is expressed in the macrophage lineage, including osteoclasts, and the CysLTR1 antagonist Montelukast has been shown to suppress the formation of osteoclasts. However, it currently remains unclear whether CysLTR1 is involved in osteoclast differentiation and bone loss. Therefore, to clarify the role of CysLTR1 in osteoclastogenesis and pathological bone loss, we herein generated CysLTR1 loss-of-function mutant mice by disrupting the cysltr1 gene using the CRISPR-Cas9 system. These mutant mice had a frameshift mutation resulting in a premature stop codon (Cysltr1 KO) or an in-frame mutation causing the deletion of the first extracellular loop (Cysltr1Δ105). Bone marrow macrophages (BMM) from these mutant mice lost the intracellular flux of calcium in response to leukotriene D4, indicating that these mutants completely lost the activity of CysLTR1 without triggering genetic compensation. However, disruption of the Cysltr1 gene did not suppress the formation of osteoclasts from BMM in vitro. We also demonstrated that the CysLTR1 antagonist Montelukast suppressed the formation of osteoclasts without functional CysLTR1. On the other hand, disruption of the Cysltr1 gene partially suppressed the formation of osteoclasts stimulated by leukotriene D4 and did not inhibit that by glutathione, functioning as a substrate in the synthesis of cysteinyl leukotrienes. Disruption of the Cysltr1 gene did not affect ovariectomy-induced osteoporosis or lipopolysaccharide-induced bone resorption. Collectively, these results suggest that the CysLT-CysLTR1 axis is dispensable for osteoclast differentiation in vitro and pathological bone loss, while the leukotriene D4-CysTR1 axis is sufficient to stimulate osteoclast formation. We concluded that the effects of glutathione and Montelukast on osteoclast formation were independent of CysLTR1.

[Two Cases at Community Pharmacies of Mercaptopurine Removal from a Disk-type Powder-packaging Machine by Wiping It with Water].

"Leukerin® powder 10%" containing mercaptopurine (6-MP) is an oral anticancer drug that requires careful handling. As a powder formulation, there are risks of exposure due to scattering during dispensing and possible 6-MP contamination to other drugs due to adhesion to the packaging machine. We previously reported that wiping with an alcohol-containing towel is useful for removing scattered powder after dispensing. However, it is recommended to wipe disk-type powder-packaging machines with water instead of cleaning with the alcohol-containing towel. Hence, we scattered 6-MP powder 100 mg (total amount of 6-MP: 10 mg), and then wiped with water three times using different types of cloth each time. We confirmed that third time wiping cloth did not have any 6-MP. Furthermore, we confirmed that the adhering 6-MP could be removed by wipe-cleaning (water-wiping twice and dry-wiping once) after dispensing 6-MP powder at two pharmacies that routinely dispensed 6-MP powder using a disk-type powder-packaging machine. In addition, we confirmed the adhesion of 6-MP in parts of the machine not cleaned by wipe-cleaning and also in parts that were washed only with water, in both the pharmacies. Based on the above observations, we recommend the following steps for cleaning disk-type powder-packaging machines after dispensing 6-MP powder: (1) wipe-cleaning that includes water-wiping twice and then dry-wiping once, (2) cleaning all areas of the packaging machine, and (3) wipe-cleaning with water before washing with water.

Molecular and Genetic Interactions between CCN2 and CCN3 behind Their Yin-Yang Collaboration.

Cellular communication network factor (CCN) 2 and 3 are the members of the CCN family that conduct the harmonized development of a variety of tissues and organs under interaction with multiple biomolecules in the microenvironment. Despite their striking structural similarities, these two members show contrastive molecular functions as well as temporospatial emergence in living tissues. Typically, CCN2 promotes cell growth, whereas CCN3 restrains it. Where CCN2 is produced, CCN3 disappears. Nevertheless, these two proteins collaborate together to execute their mission in a yin-yang fashion. The apparent functional counteractions of CCN2 and CCN3 can be ascribed to their direct molecular interaction and interference over the cofactors that are shared by the two. Recent studies have revealed the mutual negative regulation systems between CCN2 and CCN3. Moreover, the simultaneous and bidirectional regulatory system of CCN2 and CCN3 is also being clarified. It is of particular note that these regulations were found to be closely associated with glycolysis, a fundamental procedure of energy metabolism. Here, the molecular interplay and metabolic gene regulation that enable the yin-yang collaboration of CCN2 and CCN3 typically found in cartilage development/regeneration and fibrosis are described.

Comparative proteomics revealed duodenal metabolic function associated with feed efficiency in slow-growing chicken.

The Korat chicken (KR), developed in Thailand, is a slow-growing breed developed as an alternative breed for Thai chicken producers. The growing interest in slow-growing chicken meat, due to its unique taste, distinct texture, health benefits, and higher broiler welfare have led to higher market demand for KR. However, its low feed efficiency (FE) has a significant negative impact on farm profitability. Understanding the molecular mechanism regulating FE allows for designing a suitable selection program and contributing to breeding more efficient chicken for poultry production. Thus, the objective of our study was to investigate the proteome differences and possible pathways associated with FE in male KR using a label-free quantitative proteomic approach. Seventy-five KR males were individually evaluated for FE, and duodenum samples from 6 animals (3 high-FE and 3 low-FE chickens) were collected at 10 wk of age for differential abundant proteins (DAPs), protein networks, functional enrichment, and pathway analyses. In this study, we found 40 DAPs significantly associated with FE pathways, including glycolysis/gluconeogenesis, peroxisome, oxidative phosphorylation, tight junction, and cysteine and methionine metabolism. Thus, variations in observed DAPs or genes related to DAPs could be interesting biomarker candidates for selection for higher feed utilization efficiency in chicken.

Effect of cellular communication network factor 2/connective tissue growth factor on tube formation by endothelial cells derived from human periodontal ligaments.

OBJECTIVES: To clarify the role of cellular communication network factor 2/connective tissue growth factor (CCN2/CTGF) in periodontal tissue regeneration by investigating, the proliferative and tubulogenic responses of human endothelial cells obtained from the periodontal ligament to CCN2/CTGF. DESIGN: Endothelial cells were seeded on agar gel medium with or without 50 ng/mL recombinant CCN2/CTGF (rCCN2/CTGF) and cultured for 6 h. Cells were morphologically and phenotypically analyzed by immunofluorescent microscopy. A colorimetric assay was used to evaluate cell proliferation, and transmission electron microscopy (TEM) was used for ultrastructural analysis. RESULTS: The proliferation of endothelial cells was best promoted by rCCN2/CTGF at 50 ng/mL. In the control group, tube formation was not observed within 6 h. In contrast, endothelial cells seeded on the agar with 50 ng/mL rCCN2/CTGF clearly showed proliferation with network formation. Under a two-dimensional culture condition, a dense network of endothelial cells was not constructed on the plastic bottom. However, drastic morphological change was observed in the endothelial cells on the agar containing rCCN2/CTGF. The endothelial cells in the dense network were interconnected with each other and showed a tube-like structure. Tight junctions or adherens junctions were observed between the adjoining endothelial cells in the dense network. CONCLUSIONS: CCN2/CTGF was found to promote the proliferation and tubulogenesis of endothelial cells from the periodontal ligament. These results suggest that CCN2/CTGF may contribute to the regeneration of damaged periodontal tissue by activating the remaining endothelial cells.

Effect of Angiotensin II on Chondrocyte Degeneration and Protection via Differential Usage of Angiotensin II Receptors.

The renin-angiotensin system (RAS) controls not only systemic functions, such as blood pressure, but also local tissue-specific events. Previous studies have shown that angiotensin II receptor type 1 (AT1R) and type 2 (AT2R), two RAS components, are expressed in chondrocytes. However, the angiotensin II (ANG II) effects exerted through these receptors on chondrocyte metabolism are not fully understood. In this study, we investigated the effects of ANG II and AT1R blockade on chondrocyte proliferation and differentiation. Firstly, we observed that ANG II significantly suppressed cell proliferation and glycosaminoglycan content in rat chondrocytic RCS cells. Additionally, ANG II decreased CCN2, which is an anabolic factor for chondrocytes, via increased MMP9. In Agtr1a-deficient RCS cells generated by the CRISPR-Cas9 system, Ccn2 and Aggrecan (Acan) expression increased. Losartan, an AT1R antagonist, blocked the ANG II-induced decrease in CCN2 production and Acan expression in RCS cells. These findings suggest that AT1R blockade reduces ANG II-induced chondrocyte degeneration. Interestingly, AT1R-positive cells, which were localized on the surface of the articular cartilage of 7-month-old mice expanded throughout the articular cartilage with aging. These findings suggest that ANG II regulates age-related cartilage degeneration through the ANG II-AT1R axis.

Cellular communication network factor 3 in cartilage development and maintenance.

Cellular communication network factor (CCN) 3 is one of the classical members of the CCN family, which are characterized by common molecular structures and multiple functionalities. Although this protein was discovered as a gene product overexpressed in a truncated form in nephroblastoma, recent studies have revealed its physiological roles in the development and homeostasis of mammalian species, in addition to its pathological association with a number of diseases. Cartilage is a tissue that creates most of the bony parts and cartilaginous tissues that constitute the human skeleton, in which CCN3 is also differentially produced to exert its molecular missions therein. In this review article, after the summary of the molecular structure and function of CCN3, recent findings on the regulation of ccn3 expression and the roles of CCN3 in endochondral ossification, cartilage development, maintenance and disorders are introduced with an emphasis on the metabolic regulation and function of this matricellular multifunctional molecule.

Krüppel-Like Factor 4 and Its Activator APTO-253 Induce NOXA-Mediated, p53-Independent Apoptosis in Triple-Negative Breast Cancer Cells.

Inducing apoptosis is an effective treatment for cancer. Conventional cytotoxic anticancer agents induce apoptosis primarily through activation of tumor suppressor p53 by causing DNA damage and the resulting regulation of B-cell leukemia/lymphoma-2 (BCL-2) family proteins. Therefore, the effects of these agents are limited in cancers where p53 loss-of-function mutations are common, such as triple-negative breast cancer (TNBC). Here, we demonstrate that ultraviolet (UV) light-induced p53-independent transcriptional activation of NOXA, a proapoptotic factor in the BCL-2 family, results in apoptosis induction. This UV light-induced NOXA expression was triggered by extracellular signal-regulated kinase (ERK) activity. Moreover, we identified the specific UV light-inducible DNA element of the NOXA promoter and found that this sequence is responsible for transcription factor Krüppel-like factor 4 (KLF4)-mediated induction. In p53-mutated TNBC cells, inhibition of KLF4 by RNA interference reduced NOXA expression. Furthermore, treatment of TNBC cells with a KLF4-inducing small compound, APTO-253, resulted in the induction of NOXA expression and NOXA-mediated apoptosis. Therefore, our results help to clarify the molecular mechanism of DNA damage-induced apoptosis and provide support for a possible treatment method for p53-mutated cancers.

RFX1-mediated CCN3 induction that may support chondrocyte survival under starved conditions.

Cellular communication network factor (CCN) family members are multifunctional matricellular proteins that manipulate and integrate extracellular signals. In our previous studies investigating the role of CCN family members in cellular metabolism, we found three members that might be under the regulation of energy metabolism. In this study, we confirmed that CCN2 and CCN3 are the only members that are tightly regulated by glycolysis in human chondrocytic cells. Interestingly, CCN3 was induced under a variety of impaired glycolytic conditions. This CCN3 induction was also observed in two breast cancer cell lines with a distinct phenotype, suggesting a basic role of CCN3 in cellular metabolism. Reporter gene assays indicated a transcriptional regulation mediated by an enhancer in the proximal promoter region. As a result of analyses in silico, we specified regulatory factor binding to the X-box 1 (RFX1) as a candidate that mediated the transcriptional activation by impaired glycolysis. Indeed, the inhibition of glycolysis induced the expression of RFX1, and RFX1 silencing nullified the CCN3 induction by impaired glycolysis. Subsequent experiments with an anti-CCN3 antibody indicated that CCN3 supported the survival of chondrocytes under impaired glycolysis. Consistent with these findings in vitro, abundant CCN3 production by chondrocytes in the deep zones of developing epiphysial cartilage, which are located far away from the synovial fluid, was confirmed in vivo. Our present study uncovered that RFX1 is the mediator that enables CCN3 induction upon cellular starvation, which may eventually assist chondrocytes in retaining their viability, even when there is an energy supply shortage.

Risk factors for throwing elbow injuries during pitching analyzed by simulation using human musculoskeletal model in youth baseball pitchers.

BACKGROUND: Pitching mechanics are believed to be risk factors for throwing elbow injury. Thus, a prospective study of abnormal mechanics in youth baseball players is needed. This study aimed to analyze the ulnar collateral ligament during normal pitching using SIMM (Software for Interactive Musculoskeletal Modeling) for analysis and investigate the risk parameters of throwing elbow injuries in youth baseball players. We hypothesized that excessive ulnar collateral ligament force during pitching would be a risk factor for throwing elbow injuries in this population. METHODS: In this cohort study, youth baseball pitchers (aged 9-11 years) were instructed to throw a ball into a netted target. Using a SIMM musculoskeletal model, we analyzed the force of the anterior band of the anterior oblique ligament, posterior band of the anterior oblique ligament (AOL_PB), and elbow varus moment during pitching (foot contact to ball release). We calculated the integral of each force of the anterior band of the anterior oblique ligament and AOL_PB during pitching and summarized these data to establish an impulse at the medial epicondyle. Each participant was followed up for 12 months to assess the occurrence of throwing elbow injury. RESULTS: During the 12-month follow-up period, 18 pitchers (28.1%) reported throwing elbow injuries in the throwing arm. The results of this study showed that the maximum AOL_PB force and the impulse at the medial epicondyle were risk factors for throwing elbow injuries. The maximum AOL_PB force was significantly higher in the throwing elbow injury group than in the uninjured group (59.4 ± 17.8 N vs. 47.1 ± 17.5 N, P = .014). The impulse at the medial epicondyle was also significantly different (11.1 ± 4.0 N ï½¥ s in the throwing elbow injury group vs. 8.3 ± 4.4 N ï½¥ s in the uninjured group, P = .025). CONCLUSIONS: Increasing the AOL_PB force or the impulse at the medial epicondyle may increase the risk of throwing elbow injuries in youth baseball pitchers. It may be possible to reduce injury risk by focusing on ways to decrease AOL_PB load and cumulative stress on the medial epicondyle throughout the throwing motion while still maintaining high levels of ball velocity.

Therapeutic benefit of lymphadenectomy for older patients with urothelial carcinoma of the upper urinary tract: a propensity score matching study.

OBJECTIVES: Regional lymphadenectomy for urothelial carcinoma of the upper urinary tract is sometimes avoided in older patients to reduce surgical burden. We aimed to evaluate the therapeutic impact of lymphadenectomy in older patients undergoing curative therapy for upper urinary tract urothelial carcinoma. METHODS: The patients with urothelial carcinoma of the upper urinary tract older than 75 years at the time of surgery and without lymph node or distant metastasis who underwent curative therapy at two tertiary hospitals between 1994 and 2019 were retrospectively analyzed. Complete-lymphadenectomy was performed as per our protocol. Cancer-specific survival, overall survival and metastasis-free survival after surgery were evaluated between complete-lymphadenectomy and no/incomplete-lymphadenectomy groups before and after 1:1 propensity score matching. RESULTS: The original cohort included 150 patients (median age, 80.71 years), and complete-lymphadenectomy was performed in 42 (28.00%) patients. Patients in complete-lymphadenectomy group were younger and less likely to be aged >80 years (both, P < 0.0001). After matching, 30 patients were allocated to each group and the ages were comparable (78.58 vs. 77.48 years, P = 0.1738). High-grade perioperative complication rates did not differ between groups both before and after matching. Cancer-specific survival, overall survival and metastasis-free survival were significantly longer in the complete-lymphadenectomy group both before and after matching (all, P < 0.05). CONCLUSIONS: This study suggests that complete-lymphadenectomy may provide therapeutic benefits for older patients. The decision to perform complete-lymphadenectomy must be based on the patient's physical condition, rather than his/her chronological age.