A Low-Dose β1-Blocker in Combination with Milrinone Improves Intracellular Ca2+ Handling in Failing Cardiomyocytes by Inhibition of Milrinone-Induced Diastolic Ca2+ Leakage from the Sarcoplasmic Reticulum

Objectives

The purpose of this study was to investigate whether adding a low-dose β1-blocker to milrinone improves cardiac function in failing cardiomyocytes and the underlying cardioprotective mechanism.

Background

The molecular mechanism underlying how the combination of low-dose β1-blocker and milrinone affects intracellular Ca²⁺ handling in heart failure remains unclear.

Methods

We investigated the effect of milrinone plus landiolol on intracellular Ca²⁺ transient (CaT), cell shortening (CS), the frequency of diastolic Ca²⁺ sparks (CaSF), and sarcoplasmic reticulum Ca²⁺ concentration ({Ca²⁺}_SR) in normal and failing canine cardiomyocytes and used immunoblotting to determine the phosphorylation level of ryanodine receptor (RyR2) and phospholamban (PLB).

Results

In failing cardiomyocytes, CaSF significantly increased, and peak CaT and CS markedly decreased compared with normal myocytes. Administration of milrinone alone slightly increased peak CaT and CS, while CaSF greatly increased with a slight increase in {Ca²⁺}_SR. Co-administration of β1-blocker landiolol to failing cardiomyocytes at a dose that does not inhibit cardiomyocyte function significantly decreased CaSF with a further increase in {Ca²⁺}_SR, and peak CaT and CS improved compared with milrinone alone. Landiolol suppressed the hyperphosphorylation of RyR2 (Ser2808) in failing cardiomyocytes but had no effect on levels of phosphorylated PLB (Ser16 and Thr17). Low-dose landiolol significantly inhibited the alternans of CaT and CS under a fixed pacing rate (0.5 Hz) in failing cardiomyocytes.

Conclusion

A low-dose β1-blocker in combination with milrinone improved cardiac function in failing cardiomyocytes, apparently by inhibiting the phosphorylation of RyR2, not PLB, and subsequent diastolic Ca²⁺ leak.     

Turnover Rates of Hepatic Collagen and Circulating Collagen-Associated Proteins in Humans with Chronic Liver Disease

Accumulation and degradation of scar tissue in fibrotic liver disease occur slowly, typically over many years. Direct measurement of fibrogenesis, the rate of scar tissue deposition, may provide valuable therapeutic and prognostic information. We describe here results from a pilot study utilizing in vivo metabolic labeling to measure the turnover rate of hepatic collagen and collagen-associated proteins in plasma for the first time in human subjects. Eight subjects with chronic liver disease were labeled with daily oral doses of ²H₂O for up to 8 weeks prior to diagnostic liver biopsy and plasma collection. Tandem mass spectrometry was used to measure the abundance and fractional synthesis rate (FSR) of proteins in liver and blood. Relative protein abundance and FSR data in liver revealed marked differences among subjects. FSRs of hepatic type I and III collagen ranged from 0.2–0.6% per day (half-lives of 4 months to a year) and correlated significantly with worsening histologic fibrosis. Analysis of plasma protein turnover revealed two collagen-associated proteins, lumican and transforming growth factor beta-induced-protein (TGFBI), exhibiting FSRs that correlated significantly with FSRs of hepatic collagen. In summary, this is the first direct measurement of liver collagen turnover in vivo in humans and suggests a high rate of collagen remodeling in advanced fibrosis. In addition, the FSRs of collagen-associated proteins in plasma are measurable and may provide a novel strategy for monitoring hepatic fibrogenesis rates.     

Temporal and Motor Representation of Rhythm in Fronto-Parietal Cortical Areas: An fMRI Study

When sounds occur with temporally structured patterns, we can feel a rhythm. To memorize a rhythm, perception of its temporal patterns and organization of them into a hierarchically structured sequence are necessary. On the other hand, rhythm perception can often cause unintentional body movements. Thus, we hypothesized that rhythm information can be manifested in two different ways; temporal and motor representations. The motor representation depends on effectors, such as the finger or foot, whereas the temporal representation is effector-independent. We tested our hypothesis with a working memory paradigm to elucidate neuronal correlates of temporal or motor representation of rhythm and to reveal the neural networks associated with these representations. We measured brain activity by fMRI while participants memorized rhythms and reproduced them by tapping with the right finger, left finger, or foot, or by articulation. The right inferior frontal gyrus and the inferior parietal lobule exhibited significant effector-independent activations during encoding and retrieval of rhythm information, whereas the left inferior parietal lobule and supplementary motor area (SMA) showed effector-dependent activations during retrieval. These results suggest that temporal sequences of rhythm are probably represented in the right fronto-parietal network, whereas motor sequences of rhythm can be represented in the SMA-parietal network.     

Clinical Report on the First Prototype of a Photoacoustic Tomography System with Dual Illumination for Breast Cancer Imaging

Photoacoustic tomography is a recently developed imaging modality that can provide high spatial-resolution images of hemoglobin distribution in tissues such as the breast. Because breast cancer is an angiogenesis-dependent type of malignancy, we evaluated the clinical acceptability of breast tissue images produced using our first prototype photoacoustic mammography (PAM) system in patients with known cancer. Post-excisionally, histological sections of the tumors were stained immunohistochemically (IHC) for CD31 (an endothelial marker) and carbonic anhydrase IX (CAIX) (a marker of hypoxia). Whole-slide scanning and image analyses were used to evaluate the tumor microvessel distribution pattern and to calculate the total vascular perimeter (TVP)/area for each lesion. In this clinical study, 42 lesions were primarily scanned using PAM preoperatively, three of which were reported to be benign and were excluded from statistical analysis. Images were produced for 29 out of 39 cancers (visibility rate = 74.4%) at the median depth of 26.5 (3.25–51.2) mm. Age, menopausal status, body mass index, history of neoadjuvant treatment, clinical stage and histological tumor angiogenesis markers did not seem to affect the visibility. The oxygen saturation level in all of the measured lesions was lower than in the subcutaneous counterpart vessels (Wilcoxon test, p value<0.001), as well as in the counterpart contralateral normal breast region of interest (ROI) (Wilcoxon test, p value = 0.001). Although the oxygen saturation level was not statistically significant between CAIX-positive vs. -negative cases, lesional TVP/area showed a positive correlation with the oxygen saturation level only in the group that had received therapy before PAM. In conclusion, the vascular and oxygenation data obtained by PAM have great potential for identifying functional features of breast tumors.     

Cumulative Incidence and Predictors of Progression in Corticosteroid-Na?ve Patients with Sarcoidosis

Background

Assessment of the clinical course of sarcoidosis requires long-term observation. However, the appropriate period of follow-up for sarcoidosis remains unclear, especially in patients without indication of corticosteroid therapy at the time of diagnosis.

Objective

This study aimed to clarify the cumulative incidence and identify risk factors for disease progression in corticosteroid-naïve sarcoidosis patients.

Methods

The clinical courses of 150 Japanese patients with sarcoidosis, who were followed for more than 2 years and had no indication for corticosteroid therapy at diagnosis, were retrospectively reviewed. Disease progression was defined as worsening of pulmonary sarcoidosis, development of new organ involvement, or extrapulmonary organ damage. The cumulative incidence of progression was estimated by generating a cumulative incidence curve with the Fine and Gray method.

Results

The median follow-up duration was 7.7 years (interquartile range, 4.7–13.6 years). Thirty-two (21%) patients experienced disease progression. New organ involvement appeared in 16 patients (11%). The 6-month, and 1-, 5-, 10-, and 15-year cumulative incidence of progression was 2%, 5%, 15%, 28%, and 31%, respectively. The number of organs involved at diagnosis was an independent predictor for progression with a multifactorial adjusted hazard ratio of 1.71 (95% confidence interval, 1.11–2.62). The optimal cut-off of the number of organs involved at diagnosis to identify future progression was three.

Conclusions

In corticosteroid-naïve sarcoidosis patients, the risks of disease progression are comparable from 0–5 years and 5–10 years after diagnosis. The number of organs involved at diagnosis is a useful predictor for progression of sarcoidosis.     

Melatonin Inhibits Embryonic Salivary Gland Branching Morphogenesis by Regulating Both Epithelial Cell Adhesion and Morphology

Many organs, including salivary glands, lung, and kidney, are formed by epithelial branching during embryonic development. Branching morphogenesis occurs via either local outgrowths or the formation of clefts that subdivide epithelia into buds. This process is promoted by various factors, but the mechanism of branching morphogenesis is not fully understood. Here we have defined melatonin as a potential negative regulator or “brake” of branching morphogenesis, shown that the levels of it and its receptors decline when branching morphogenesis begins, and identified the process that it regulates. Melatonin has various physiological functions, including circadian rhythm regulation, free-radical scavenging, and gonadal development. Furthermore, melatonin is present in saliva and may have an important physiological role in the oral cavity. In this study, we found that the melatonin receptor is highly expressed on the acinar epithelium of the embryonic submandibular gland. We also found that exogenous melatonin reduces salivary gland size and inhibits branching morphogenesis. We suggest that this inhibition does not depend on changes in either proliferation or apoptosis, but rather relates to changes in epithelial cell adhesion and morphology. In summary, we have demonstrated a novel function of melatonin in organ formation during embryonic development.     

Baseline Assessment of Mesophotic Reefs of the Vitória-Trindade Seamount Chain Based on Water Quality,Microbial Diversity,Benthic Cover and Fish Biomass Data

Seamounts are considered important sources of biodiversity and minerals. However, their biodiversity and health status are not well understood; therefore, potential conservation problems are unknown. The mesophotic reefs of the Vitória-Trindade Seamount Chain (VTC) were investigated via benthic community and fish surveys, metagenomic and water chemistry analyses, and water microbial abundance estimations. The VTC is a mosaic of reef systems and includes fleshy algae dominated rhodolith beds, crustose coralline algae (CCA) reefs, and turf algae dominated rocky reefs of varying health levels. Macro-carnivores and larger fish presented higher biomass at the CCA reefs (4.4 kg per frame) than in the rhodolith beds and rocky reefs (0.0 to 0.1 kg per frame). A larger number of metagenomic sequences identified as primary producers (e.g., Chlorophyta and Streptophyta) were found at the CCA reefs. However, the rocky reefs contained more diseased corals (>90%) than the CCA reefs (~40%) and rhodolith beds (~10%). Metagenomic analyses indicated a heterotrophic and fast-growing microbiome in rocky reef corals that may possibly lead to unhealthy conditions possibly enhanced by environmental features (e.g. light stress and high loads of labile dissolved organic carbon). VTC mounts represent important hotspots of biodiversity that deserve further conservation actions.     

Angiotensin II Reduces Lipoprotein Lipase Expression in Visceral Adipose Tissue via Phospholipase C β4 Depending on Feeding but Increases Lipoprotein Lipase Expression in Subcutaneous Adipose Tissue via c-Src

Metabolic syndrome is characterized by visceral adiposity, insulin resistance, high triglyceride (TG)- and low high-density lipoprotein cholesterol-levels, hypertension, and diabetes—all of which often cause cardiovascular and cerebrovascular diseases. It remains unclear, however, why visceral adiposity but not subcutaneous adiposity causes insulin resistance and other pathological situations. Lipoprotein lipase (LPL) catalyzes hydrolysis of TG in plasma lipoproteins. In the present study, we investigated whether the effects of angiotensin II (AngII) on TG metabolism are mediated through an effect on LPL expression. Adipose tissues were divided into visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) for comparison. AngII accelerated LPL expression in SAT but, on the contrary, suppressed its expression in VAT. In both SAT and VAT, AngII signaled through the same type 1 receptor. In SAT, AngII increased LPL expression via c-Src and p38 MAPK signaling. In VAT, however, AngII reduced LPL expression via the G_q class of G proteins and the subsequent phospholipase C β4 (PLCβ4), protein kinase C β1, nuclear factor κB, and inducible nitric oxide synthase signaling pathways. PLCβ4 small interfering RNA experiments showed that PLCβ4 expression is important for the AngII-induced LPL reduction in VAT, in which PLCβ4 expression increases in the evening and falls at night. Interestingly, PLCβ4 expression in VAT decreased with fasting, while AngII did not decrease LPL expression in VAT in a fasting state. In conclusion, AngII reduces LPL expression through PLCβ4, the expression of which is regulated by feeding in VAT, whereas AngII increases LPL expression in SAT. The different effects of AngII on LPL expression and, hence, TG metabolism in VAT and SAT may partly explain their different contributions to the development of metabolic syndrome.