Uric Acid Spherulites in the Reflector Layer of Firefly Light Organ

Background

In firefly light organs, reflector layer is a specialized tissue which is believed to play a key role for increasing the bioluminescence intensity through reflection. However, the nature of this unique tissue remains elusive. In this report, we investigated the role, fine structure and nature of the reflector layer in the light organ of adult Luciola cerata.

Principal Findings

Our results indicated that the reflector layer is capable of reflecting bioluminescence, and contains abundant uric acid. Electron microscopy (EM) demonstrated that the cytosol of the reflector layer''s cells is filled with densely packed spherical granules, which should be the uric acid granules. These granules are highly regular in size (∼700 nm in diameter), and exhibit a radial internal structure. X-ray diffraction (XRD) analyses revealed that an intense single peak pattern with a d-spacing value of 0.320 nm is specifically detected in the light organ, and is highly similar to the diffraction peak pattern and d-spacing value of needle-formed crystals of monosodium urate monohydrate. However, the molar ratio evaluation of uric acid to various cations (K⁺, Na⁺, Ca²⁺ and Mg²⁺) in the light organ deduced that only a few uric acid molecules were in the form of urate salts. Thus, non-salt uric acid should be the source of the diffraction signal detected in the light organ.

Conclusions

In the light organ, the intense single peak diffraction signal might come from a unique needle-like uric acid form, which is different from other known structures of non-salt uric acid form. The finding of a radial structure in the granules of reflector layer implies that the spherical uric acid granules might be formed by the radial arrangement of needle-formed packing matter.     

HER2 Amplification and Clinicopathological Characteristics in a Large Asian Cohort of Rare Mucinous Ovarian Cancer

Mucinous epithelial ovarian cancer has a poor prognosis in the advanced stages and responds poorly to conventional chemotherapy. We aim to elucidate the clinicopathological factors and incidence of HER2 expression of this cancer in a large Asian retrospective cohort from Singapore. Of a total of 133 cases, the median age at diagnosis was 48.3 years (range, 15.8–89.0 years), comparatively younger than western cohorts. Most were Chinese (71%), followed by Malays (16%), others (9.0%), and Indians (5%). 24% were noted to have a significant family history of malignancy of which breast and gastrointestinal cancers the most prominent. Majority of the patients (80%) had stage I disease at diagnosis. Information on HER2 status was available in 113 cases (85%). Of these, 31 cases (27.4%) were HER2+, higher than 18.8% reported in western population. HER2 positivity appeared to be lower among Chinese and higher among Malays patients (p = 0.052). With the current standard of care, there was no discernible impact of HER2 status on overall survival. (HR = 1.79; 95% CI, 0.66–4.85; p = 0.249). On the other hand, positive family history of cancer, presence of lymphovascular invasion, and ovarian surface involvements were significantly associated with inferior overall survival on univariate and continued to be statistically significant after adjustment for stage. While these clinical factors identify high risk patients, it is promising that the finding of a high incidence of HER2 in our Asian population may allow development of a HER2 targeted therapy to improve the management of mucinous ovarian cancers.     

Stress transfer in collagen fibrils reinforcing connective tissues: effects of collagen fibril slenderness and relative stiffness

Unlike engineering fibre composite materials which comprise of fibres that are uniform cylindrical in shape, collagen fibrils reinforcing the proteoglycan-rich (PG) gel in the extra-cellular matrices (ECMs) of connective tissues are taper-ended (paraboloidal in shape). In an earlier paper we have discussed how taper of a fibril leads to an axial stress up-take which differs from that of a uniform cylindrical fibre and implications for fibril fracture. The present paper focuses on the influence of fibre aspect ratio, q (slenderness), and Young's modulus (stiffness), relative to that of the gel phase, E(R), on the magnitude of the axial tensile stresses generated within a fibril and wider implications on failure at tissue level. Fibre composite models were evaluated using finite element (FE) and mathematical analyses. When the applied force is low, there is elastic stress transfer between the PG gel and a fibril. FE modelling shows that the stress in a fibril increases with E(R) and q. At higher applied forces, there is plastic stress transfer. Mathematical modelling predicts that the stress in a fibril increases linearly with q. For small q values, fibrils may be regarded as fillers with little ability to provide tensile reinforcement. Large q values lead to high stress in a fibril. Such high stresses are beneficial provided they do not exceed the fracture stress of collagen. Modulus difference regulates the strain energy release density, u, for interfacial rupture; large E(R) not only leads to high stress in a fibril but also insures against interfacial rupture by raising the value of u.     

Elicitor-induced cytoskeletal rearrangement relates to vacuolar dynamics and execution of cell death: in vivo imaging of hypersensitive cell death in tobacco BY-2 cells

Disintegration of the vacuolar membrane (VM) has been proposed to be a crucial event in various types of programmed cell death (PCD) in plants. However, its regulatory mechanisms are mostly unknown. To obtain new insights on the regulation of VM disintegration during hypersensitive cell death, we investigated the structural dynamics and permeability of the VM, as well as cytoskeletal reorganization during PCD in tobacco BY-2 cells induced by a proteinaceous elicitor, cryptogein. From sequential observations, we have identified the following remarkable events during PCD. Stage 1: bulb-like VM structures appear within the vacuolar lumen and the cortical microtubules are disrupted, while the cortical actin microfilaments are bundled. Simultaneously, transvacuolar strands including endoplasmic microtubules and actin microfilaments are gradually disrupted and the nucleus moves from the center to the periphery of the cell. Stage 2: cortical actin microfilament bundles and complex bulb-like VM structures disappear. The structure of the large central vacuole becomes simpler, and small spherical vacuoles appear. Stage 3: the VM is disintegrated and a fluorescent dye, BCECF, leaks out of the vacuoles just prior to PCD. Application of an actin polymerization inhibitor facilitates both the disappearance of bulb-like vacuolar membrane structures and induction of cell death. These results suggest that the elicitor-induced reorganization of actin microfilaments is involved in the regulation of hypersensitive cell death via modification of the vacuolar structure to induce VM disintegration.     

A 12-week aerobic exercise intervention results in improved metabolic function and lower adipose tissue and ectopic fat in high-fat diet fed rats

Investigations of long-term exercise interventions in humans to reverse obesity is expensive and is hampered by poor compliance and confounders. In the present study, we investigated intrahepatic and muscle fat, visceral and subcutaneous fat pads, plasma metabolic profile and skeletal muscle inflammatory markers in response to 12-week aerobic exercise in an obese rodent model. Six-week-old male Wistar rats (n=20) were randomized to chow-fed control (Control, n=5), sedentary high-fat diet (HFD, n=5), chow-fed exercise (Exercise, n=5) and HFD-fed exercise (HFD+Exercise, n=5) groups. The exercise groups were subjected to 12 weeks of motorized treadmill running at a speed of 18 m/min for 30 min/day. Differences in post-intervention measures were assessed by analysis of covariance (ANCOVA), adjusted for baseline bodyweight and pre-intervention measures, where available. Post-hoc analyses were performed with Bonferroni correction. Plasma metabolic profile was worsened and fat pads, ectopic fat in muscle and liver and inflammatory markers in skeletal muscle were elevated in sedentary HFD-fed animals relative to chow-fed controls. HFD+Exercise animals had significantly lower leptin (P=0.0004), triglycerides (P=0.007), homeostatic model assessment of insulin resistance (HOMA-IR; P=0.065), intramyocellular lipids (IMCLs; P=0.003), intrahepatic lipids (IHLs; P<0.0001), body fat% (P=0.001), subcutaneous adipose tissue (SAT; P<0.0001), visceral adipose (P<0.0001) and total fat mass (P<0.0001), relative to sedentary HFD-fed animals, despite only modestly lower bodyweight. Messenger RNA (mRNA) expression of inflammatory markers Interleukin 6 (IL6) and Tumor necrosis factor α (TNFα) were also reduced with aerobic exercise in skeletal muscle. Our results suggest that 12 weeks of aerobic exercise training is effective in improving metabolic health, fat depots, ectopic fat and inflammation even against a high-fat dietary background.     

A thermomechanical framework for reconciling the effects of ultraviolet radiation exposure time and wavelength on connective tissue elasticity

Augmentation of the mechanical properties of connective tissue using ultraviolet (UV) radiation—by targeting collagen cross-linking in the tissue at predetermined UV exposure time \((t)\) and wavelength \((\lambda )\) —has been proposed as a therapeutic method for supporting the treatment for structural-related injuries and pathologies. However, the effects of \(\lambda \) and \(t\) on the tissue elasticity, namely elastic modulus \((E)\) and modulus of resilience \((u_\mathrm{Y})\) , are not entirely clear. We present a thermomechanical framework to reconcile the \(t\) - and \(\lambda \) -related effects on \(E\) and \(u_\mathrm{Y}\) . The framework addresses (1) an energy transfer model to describe the dependence of the absorbed UV photon energy, \(\xi \) , per unit mass of the tissue on \(t\) and \(\lambda \) , (2) an intervening thermodynamic shear-related parameter, \(G\) , to quantify the extent of UV-induced cross-linking in the tissue, (3) a threshold model for the \(G\) versus \(\xi \) relationship, characterized by   \(t_\mathrm{C}\) —the critical \(t\) underpinning the association of \(\xi \) with \(G\) —and (4) the role of \(G\) in the tissue elasticity. We hypothesized that \(G\) regulates \(E\) (UV-stiffening hypothesis) and \(u_\mathrm{Y}\) (UV-resilience hypothesis). The framework was evaluated with the support from data derived from tensile testing on isolated ligament fascicles, treated with two levels of \(\lambda \) (365 and 254 nm) and three levels of \(t\) (15, 30 and 60 min). Predictions from the energy transfer model corroborated the findings from a two-factor analysis of variance of the effects of \(t\) and \(\lambda \) treatments. Student’s t test revealed positive change in \(E\) and \(u_\mathrm{Y}\) with increases in \(G\) —the findings lend support to the hypotheses, implicating the implicit dependence of UV-induced cross-links on \(t\) and \(\lambda \) for directing tissue stiffness and resilience. From a practical perspective, the study is a step in the direction to establish a UV irradiation treatment protocol for effective control of exogenous cross-linking in connective tissues.     

Predicting Survival of De Novo Metastatic Breast Cancer in Asian Women: Systematic Review and Validation Study

Background

In Asia, up to 25% of breast cancer patients present with distant metastases at diagnosis. Given the heterogeneous survival probabilities of de novo metastatic breast cancer, individual outcome prediction is challenging. The aim of the study is to identify existing prognostic models for patients with de novo metastatic breast cancer and validate them in Asia.

Materials and Methods

We performed a systematic review to identify prediction models for metastatic breast cancer. Models were validated in 642 women with de novo metastatic breast cancer registered between 2000 and 2010 in the Singapore Malaysia Hospital Based Breast Cancer Registry. Survival curves for low, intermediate and high-risk groups according to each prognostic score were compared by log-rank test and discrimination of the models was assessed by concordance statistic (C-statistic).

Results

We identified 16 prediction models, seven of which were for patients with brain metastases only. Performance status, estrogen receptor status, metastatic site(s) and disease-free interval were the most common predictors. We were able to validate nine prediction models. The capacity of the models to discriminate between poor and good survivors varied from poor to fair with C-statistics ranging from 0.50 (95% CI, 0.48–0.53) to 0.63 (95% CI, 0.60–0.66).

Conclusion

The discriminatory performance of existing prediction models for de novo metastatic breast cancer in Asia is modest. Development of an Asian-specific prediction model is needed to improve prognostication and guide decision making.     

Investigation of differentially expressed proteins due to the inhibitory effects of berberine in human liver cancer cell line HepG2

The investigation of differentially expressed proteins was used together with other techniques to study the inhibitory effects of two different doses of berberine in human liver cancer cell line HepG2. For HepG2 cells treated with 24.0 mg l(-1) of berberine, an increase in the sub G(0) phase that was indicative of cell death was observed in cell cycle analysis with flow cytometry. However, no significant increase in sub G(0) was observed in HepG2 cells treated with 4.0 mg l(-1) of berberine. Using flow cytometric analysis, significant activation of caspase 3 was not observed with HepG2 cells treated with 4.0 and 24.0 mg l(-1) of berberine. In this work, labeling of cells with stable isotope was not used in the proposed method developed. The whole cell lysates from the control and treated cells were digested with trypsin and the peptides were separated by two-dimensional (cation exchange and reversed phase) liquid chromatography and tandem mass spectrometry. Our preliminary data showed that the proposed platform provided a rapid approach to study the molecular mechanism due to the inhibitory effects of different doses of the berberine on HepG2 cell lines. This included a network of proteins involved in mitogen-activated protein kinase (MAPK) phosphorelay systems, metabolism, regulation of cell cycle and DNA damage response. The differentially expressed proteins identified using the current approach were consistent with the data obtained from cell cycle analysis with flow cytometry.