Ischemia-reperfusion (IR) of the testis results in germ cell-specific apoptosis and can lead to aspermatogenesis. Germ cell-specific apoptosis after IR of the testis has been shown to be correlated with and dependent on neutrophil recruitment to the testis after IR. Studies that used E-selectin-deficient mice have demonstrated that E-selectin expression is critical for neutrophil recruitment to subtunical venules in the testis after IR and for the resultant germ cell-specific apoptosis. The present study investigates the in vivo signaling pathway that exists after IR that leads to neutrophil recruitment in the murine testis. Mice were subjected to a 2-h period of testicular ischemia followed by reperfusion. Results demonstrate that the proinflammatory cytokines, tumor necrosis factor alpha (TNFalpha) and interleukin 1beta (IL-1beta), are stimulated after IR as is the phosphorylation of c-jun N-terminal kinase (JNK). The downstream transcription factors of JNK, ATF-2 and c-jun are also phosphorylated at specific times after IR of the testis. Activation of the JNK stress-related kinase pathway is correlated with an increase in E-selectin expression and neutrophil recruitment to the testis after IR. Intratesticular injection of IL-1beta also caused JNK phosphorylation and neutrophil recruitment to the testis. These results suggest that testicular IR injury stimulates IL-1beta expression, which leads to activation of the JNK signaling pathway and ultimately E-selectin expression and neutrophil recruitment to the testis. This provides the first evidence of a cytokine/stress-related kinase signaling pathway to E-selectin expression in vivo.

Objective Little is known regarding the role(s) B cells play in obesity-induced metabolic dysfunction. The present study utilized a mouse with B cell-specific deletion of Id3 (Id3Bcell KO) to identify B cell functions involved in the metabolic consequences of obesity. Approach and Results Diet-induced obese (DIO) Id3Bcell KO mice demonstrated attenuated inflammation and insulin resistance in visceral adipose tissue (VAT), and improved systemic glucose tolerance. VAT in Id3Bcell KO mice had increased B-1b B cells and elevated IgM natural antibodies (Nabs) to oxidation-specific epitopes (OSE). B-1b B cells reduced cytokine production in VAT M1 macrophages, and adoptively transferred B-1b B cells trafficked to VAT and produced NAbs for the duration of thirteen week studies. B-1b B cells null for Id3 demonstrated increased proliferation, established larger populations in Rag1−/− VAT, and attenuated diet-induced glucose intolerance and VAT insulin resistance in Rag1−/− hosts. However, transfer of B-1b B cells unable to secrete IgM had no effect on glucose tolerance. In an obese human population, results provided the first evidence that B-1 cells are enriched in human VAT and IgM antibodies to OSEs inversely correlated with inflammation and insulin resistance. Conclusions Nab-producing B-1b B cells are increased in Id3Bcell KO mice and attenuate adipose tissue inflammation and glucose intolerance in DIO mice. Additional findings are the first to identify VAT as a reservoir for human B-1 cells and to link anti-inflammatory IgM antibodies with reduced inflammation and improved metabolic phenotype in obese humans.

Objective Inhibitor of differentiation-3 (Id3) has been implicated in promoting angiogenesis, a key determinant of high fat diet (HFD)-induced visceral adiposity. Yet the role of Id3 in high fat diet (HFD)-induced angiogenesis and visceral adipose expansion is unknown. Methods and Results Id3−/− mice demonstrated a significant attenuation of HFD-induced visceral fat depot expansion compared to WT littermate controls. Importantly, unlike other Id proteins, loss of Id3 did not affect adipose depot size in young mice fed chow diet or differentiation of adipocytes in vitro or in vivo. Contrast enhanced ultrasound revealed a significant attenuation of visceral fat microvascular blood volume in HFD-fed mice null for Id3 compared to WT controls. HFD induced Id3 and VEGFA expression in the visceral stromal vascular fraction (SVF) and Id3−/− mice had significantly lower levels of VEGFA protein in visceral adipose tissue compared to WT. Furthermore, HFD-induced VEGFA expression in visceral adipose tissue was completely abolished by loss of Id3. Consistent with this effect, Id3 abolished E12-mediated repression of VEGFA promoter activity. Conclusions Results identify Id3 as an important regulator of HFD-induced visceral adipose VEGFA expression, microvascular blood volume, and depot expansion. Inhibition of Id3 may have potential as a therapeutic strategy to limit visceral adiposity.

Hypoxia-inducible factor-1a (HIF-1a) is a transcription factor that plays an essential role in oxygen homeostasis. HIF-1a is constitutively made in cells; however, it is ubiquitinated and degraded under normoxic conditions. Hypoxia prevents the ubiquitination of HIF-1a, resulting in stabilization of the protein and activation of target genes. Because of its vascular arrangement and the high metabolic demand of spermatogenesis, the testis has been described previously as functioning on the brink of hypoxia; thus, we have hypothesized that HIF-1a is constitutively expressed and stabilized in the testis, where it could play a role in testicular homeostasis. Western blot analysis using nuclear proteins from liver, kidney, and testis revealed the presence of HIF-1a only in the testis. Immunohistochemistry confirmed this result and revealed that HIF1a was specifically located in interstitial Leydig cells. Electromobility shift assays employing nuclear extracts from the TM3 Leydig cell line revealed that these cells express HIF-1a that is capable of binding DNA under normoxic conditions. Furthermore, we found that protein levels can be increased further when the TM3 cells are cultured under hypoxic conditions. Finally, transient transfections of TM3 Leydig cells revealed that the promoter of the mouse 3b-hydroxysteroid dehydrogenase type 1 (Hsd3b1) gene, which encodes a key enzyme in testosterone production, is a potential target of HIF-1a. In conclusion, HIF-1a is constitutively present in the Leydig cells of the murine testis, where it potentially regulates Hsd3b1 transcription, and thus male reproductive function.

Background Hypoglycemia is a known risk of intensive postoperative glucose control in cardiac surgery patients. However, neither the consequences of hypoglycemia relative to hyperglycemia, nor the possible interaction effects, have been well described. We examined the effects of postoperative hypoglycemia, hyperglycemia, and their interaction on short-term morbidity and mortality. Methods Single institution Society of Thoracic Surgeons (STS) database patient records from 2010–2014 were merged with clinical data, including blood glucose values measured in the intensive care unit (ICU). Exclusion criteria included fewer than 3 glucose measurements and absence of a STS predicted risk of morbidity or mortality score. Primary outcomes were operative mortality and composite major morbidity (permanent stroke, renal failure, prolonged ventilation, pneumonia or myocardial infarction). Secondary outcomes included ICU and postoperative length of stay. Hypoglycemia was defined as <70 mg/dL, and hyperglycemia as >180 mg/dL. Simple and multivariable regression models were used to evaluate outcomes. Results A total of 2,285 patient records met selection criteria for analysis. The mean postoperative glucose was 140.8 ± 18.8 mg/dL. Overall, 21.4% of patients experienced a hypoglycemic episode (n=488), and 1.05% (n=24) had a severe hypoglycemic episode (<40 mg/dL). The unadjusted odds ratio (UOR) for operative mortality for patients with any hypoglycemic episode compared to those without was 5.47 (95% CI 3.14–9.54), and the UOR for major morbidity was 4.66 (95% CI 3.55–6.11). After adjustment for predicted risk of morbidity or mortality and other significant covariates, the adjusted odds (AOR) of operative mortality are significant for patients with any hypoglycemia (AOR 4.88, 95% CI 2.67–8.92) and patients with both events (AOR 8.29 95% CI 1.83–37.5) but not hyperglycemia alone (AOR 1.62, 95% CI 0.56–4.69). The AOR of major morbidity for patients with both hypo- and hyperglycemic events is 14.3 (95% CI 6.50–31.4). Conclusions Postoperative hypoglycemia is associated with both mortality and major morbidity after cardiac surgery. The combination of both hyperglycemia and hypoglycemia represents a substantial increase in risk. Although it remains unclear whether hypoglycemia is a cause, an early warning sign, or a result of adverse events, this study suggests hypoglycemia may be an important event in the postoperative period after cardiac surgery.