The inhibitory effect on aldose reductase by an extract of Ganoderma lucidum

The human aldose reductase inhibitory effects of the methanol extracts of 17 medicinal and edible mushrooms were examined. Ganoderma lucidum showed the highest aldose reductase inhibitory activity compared with the other mushrooms. The effect of an ethanol extract of G. lucidum on the galactitol level in the eye lens was studied in a galactosemic rat model in vivo. This mushroom significantly decreased the galactitol accumulation. Copyright © 2008 John Wiley & Sons, Ltd.     

Clostridium botulinum type E toxins bind to Caco-2 cells by a different mechanism from that of type A toxins

Cultured Clostridium botulinum strains produce progenitor toxins designated as 12S, 16S, and 19S toxins. The 12S toxin consists of a neurotoxin (NTX, 7S) and a non-toxic non-hemagglutinin (NTNH). The 16S and 19S toxins are formed by conjugation of the 12S toxin with hemagglutinin (HA), and the 19S toxin is a dimer of the 16S toxin. Type A cultures produce all 3 of these progenitor toxins, while type E produces only the 12S toxin. The 7S toxin is cleaved into heavy (H) and light (L) chains by a protease(s) in some strains, and the H chain has 2 domains, the N-terminus (Hn) and C-terminus (Hc). It has been reported that type A toxins bind to the intestinal cells or cultured cells via either HA or Hc. In this study, we investigated the binding of type A and E toxins to Caco-2 cells using Western blot analysis. Both the type E 7S and 12S toxins bound to the cells, with the 7S toxin binding more strongly, whereas, in the type A strain, only the 16S/19S toxins showed obvious binding. Pre-incubation of the type E 7S toxin with IgG against recombinant type E Hc significantly inhibited the 7S toxin binding, indicating that Hc might be a main binding domain of the type E toxin.     

Breast cancer therapy affects the expression of antineonatal Nav1.5 antibodies in the serum of patients with breast cancer

Neonatal Nav1.5 (nNav1.5) is the alternative splice variant of Nav1.5 and it has been widely associated with the progression of breast cancer. The immunological context of nNav1.5 with respect to breast cancer metastases remains unexplored. The presence of antibodies against nNav1.5 may highlight the immunogenicity of nNav1.5. Hence, the aim of the present study was to detect the presence of antineonatal Nav1.5 antibodies (antinNav1.5-Ab) in the serum of patients with breast cancer and to elucidate the effects of breast cancer therapy on its expression. A total of 32 healthy female volunteers and 64 patients with breast cancer were randomly recruited into the present study as the control and breast cancer group, respectively. Patients with breast cancer were divided equally based on their pre- and ongoing-treatment status. Serum samples were tested with in-house indirect enzyme-linked immunosorbent assay (ELISA) to detect antinNav1.5-Ab, CD25 (T regulatory cell marker) using an ELISA kit and Luminex assay to detect the expression of metastasis-associated cytokines, such as vascular endothelial growth factor (VEGF), interleukin (IL)-6, IL-10, IL-8, chemokine (C-C motif) ligand 2 and tumor necrosis factor-alpha (TNF-α) The mean difference in the expression of antinNav1.5-Ab among the three groups (control, pretreatment and ongoing-treatment) was significant (P=0.0005) and the pretreatment breast cancer group exhibited the highest expression. The concentration of CD25 was highest in the pretreatment breast cancer group compared with the control and ongoing-treatment groups. There was a significant positive correlation between antinNav1.5-Ab and IL-6 in the pretreatment group (r=0.7260; P=0.0210) and a significant negative correlation between antinNav1.5-Ab and VEGF in the ongoing-treatment group (r=−0.842; P-value=0.0040). The high expression of antinNav1.5-Ab in the pretreatment group was in accordance with the uninterrupted presence of metastasis and highlighted the immunogenicity of nNav1.5 whereas the low expression of antinNav1.5-Ab in the ongoing-treatment group reflected the efficacy of breast cancer therapy in eliminating metastases. The augmented manifestation of T regulatory cells in the pretreatment group highlighted the functional role of nNav1.5 in promoting metastasis. The parallel expression of antinNav1.5-Ab with the imbalanced expression of cytokines promoting metastasis (IL-8, IL-6 and TNF-α) and cytokines that prevent metastasis (IL-10) indicated the role of nNav1.5 in breast cancer growth. The expression of antinNav1.5-Ab in accordance to the metastatic microenvironment indicates the immunogenicity of the protein and highlights the influence of breast cancer therapy on its expression level.     

Humoral immune responses to Epstein-Barr virus encoded tumor associated proteins and their putative extracellular domains in nasopharyngeal carcinoma patients and regional controls

Epstein-Barr virus (EBV) latency proteins EBNA1, LMP1, LMP2, and BARF1 are expressed in tumor cells of nasopharyngeal carcinoma (NPC). IgG and IgA antibody responses to these non-self tumor antigens were analyzed in NPC patients (n=125) and regional controls (n=100) by three approaches, focusing on the putative LMP1, LMP2 extracellular domains. Despite abundant IgG and IgA antibody responses to lytic antigens and EBNA1, patients had low titer (1:25-1:100) IgG to LMP1 (81.2%), LMP2 (95.6%), and BARF1 (84.8%), while immunoblot showed such reactivity in 24.2%, 12.5%, and 12.5% at 1:50 dilution, respectively. Few IgA responses were detected, except for EBNA1. Controls only showed IgG to EBNA1. ELISA using peptides from different domains of LMP1, LMP2, and BARF1 also yielded mostly negative results. When existing, low level IgG to intracellular C-terminus of LMP1 (62.9%) prevailed. Rabbit immunization with peptides representing extracellular (loop) domains yielded loop-specific antibodies serving as positive control. Importantly, these rabbit antibodies stained specifically extracellular domains of LMP1 and LMP2 on viable cells and mediated complement-driven cytolysis. Rabbit anti-LMP1 loop-1 and -3 killed 50.4% and 59.4% of X50/7 and 35.0% and 35.9% of RAJI cells, respectively, and 22% of both lines were lysed by anti-LMP2 loop-2 or -5 antibodies. This demonstrates that (extracellular domains of) EBV-encoded tumor antigens are marginally immunogenic for humoral immune responses. However, peptide-specific immunization may generate such antibodies, which can mediate cell killing via complement activation. This opens options for peptide-based tumor vaccination in patients carrying EBV latency type II tumors such as NPC.     

The combined treatment of brassinin and imatinib synergistically downregulated the expression of MMP‐9 in SW480 colon cancer cells

In cancer treatment, which is a major cause of mortality today, combination studies with clinically used chemotherapeutics are becoming increasingly important as much as investigating the effects of novel natural compounds. In this context, phytoalexins constitute an important group due to their unique structure. Brassinin is an essential indole phytoalexin and is a biosynthetic precursor for other phytoalexins. The purpose of this study was to evaluate the anticancer effects of brassinin in combination with imatinib in SW480 cells. In the study, it was observed that brassinin–imatinib combination significantly increased cytotoxicity compared with the single treatment of both compounds and inhibited cell cycle at G0/G1 phase. Annexin V binding and fluorescence imaging assays showed that the combination of brasinin–imatinib induces apoptosis in a dose‐dependent manner. Furthermore, the effect of brassinin on the activity of MMP‐9 in SW480 cells was evaluated for the first time, and it was detected that MMP‐9 activity was significantly reduced. The combination of brassinin–imatinib was found to inhibit MMP‐9 activity as well as relative MMP‐9 gene expression on a higher level compared with control and compounds alone. Our findings have revealed that the combination of brassinin–imatinib synergistically induces cytotoxicity and apoptosis in SW480 cells. The findings on MMP‐9 downregulation have also revealed the anti‐metastatic potential of treatment.     

Vitamin B12 protects against DNA damage induced by hydrochlorothiazide

DNA damage induced by hydrochlorothiazide was previously reported in cultured human lymphocytes. In this study, we aimed to investigate the harmful effects of hydrochlorothiazide on DNA by measuring the spontaneous frequency of sister chromatid exchanges (SCEs) in cultured human lymphocytes. We also aimed to investigate the possible protection of that damage by vitamin B12. The results showed that hydrochlorothiazide (5?µg/mL) significantly increased the frequency of sister chromatid exchanges (P?<?0.001) in human lymphocytes in comparison with control. Additionally, the frequency of hydrochlorothiazide-induced SCEs was significantly decreased by co-treatment with vitamin B12 at concentration of 13.5?µg/mL (P?<?0.001). In conclusion, hydrochlorothiazide is genotoxic to human lymphocytes and its toxicity is reduced by vitamin B12.     

Lung adenocarcinoma with mixed solid and crazy‐paving appearances

“Crazy paving” on CT images refers to the superimposition of ground‐glass opacity and linear pattern resembling irregularly shaped paving stones. Initially, “crazy paving” was described as a pathognomonic sign of alveolar proteinosis. Subsequently, this pattern has been reported in a variety of disorders of the lung. We demonstrated CT images of lung adenocarcinoma demonstrating both solid and crazy‐paving appearances.     

Invasion and Metastasis Suppression by Anti-Neonatal Nav1.5 Antibodies in Breast Cancer

Background: Detectable neonatal Nav1.5 (nNav1.5) expression in tumour breast tissue positive for lymph node metastasis and triple-negative subtype serves as a valid tumour-associated antigen to target and prevent breast cancer invasion and metastasis. Therapeutic antibodies against tumour antigens have become the predominant class of new drugs in cancer therapy because of their fewer adverse effects and high specificity. Objective: This study was designed to investigate the therapeutic and anti-metastatic potential of the two newly obtained anti-nNav1.5 antibodies, polyclonal anti-nNav1.5 (pAb-nNav1.5) and monoclonal anti-nNav1.5 (mAb-nNav1.5), on breast cancer invasion and metastasis. Methods: MDA-MB-231 and 4T1 cells were used as in vitro models to study the effect of pAb-nNav1.5 (59.2 µg/ml) and mAb-nNav1.5 (10 µg/ml) (24 hours treatment) on cell invasion. 4T1-induced mammary tumours in BALB/c female mice were used as an in vivo model to study the effect of a single dose of intravenous pAb-nNav1.5 (1 mg/ml) and mAb-nNav1.5 (1 mg/ml) on the occurrence of metastasis. Real-time PCR and immunofluorescence staining were conducted to assess the effect of antibody treatment on nNav1.5 mRNA and protein expression, respectively. The animals’ body weight, organs, lesions, and tumour mass were also measured and compared. Results: pAb-nNav1.5 and mAb-nNav1.5 treatments effectively suppressed the invasion of MDA-MB-231 and 4T1 cells in the 3D spheroid invasion assay. Both antibodies significantly reduced nNav1.5 gene and protein expression in these cell lines. Treatment with pAb-nNav1.5 and mAb-nNav1.5 successfully reduced mammary tumour tissue size and mass and prevented lesions in vital organs of the mammary tumour animal model whilst maintaining the animal’s healthy weight. mRNA expression of nNav1.5 in mammary tumour tissues was only reduced by mAb-nNav1.5. Conclusion: Overall, this work verifies the uniqueness of targeting nNav1.5 in breast cancer invasion and metastasis prevention, but more importantly, humanised versions of mAb-nNav1.5 may be valuable passive immunotherapeutic agents to target nNav1.5 in breast cancer.     

Neuroprotection by trans-resveratrol against collagenase-induced neurological and neurobehavioural deficits in rats involves adenosine A1 receptors

ABSTRACT

Objective: Trans-resveratrol has been shown to have neuroprotective effects and could be a promising therapeutic agent in the treatment of intracerebral haemorrhage (ICH). This study aimed to investigate the involvement of the adenosine A1 receptor (A1R) in trans-resveratrol-induced neuroprotection in rats with collagenase-induced ICH.

Methods: Sixty male Sprague–Dawley rats weighing 330–380 g were randomly divided into five groups (n = 12): (i) control, (ii) sham-operated rats, (iii) ICH rats pretreated with vehicle (0.1% DMSO saline, i.c.v.), (iv) ICH rats pretreated with trans-resveratrol (0.9 µg, i.c.v.) and (v) ICH rats pretreated with trans-resveratrol (0.9 µg) and the A1R antagonist, DPCPX (2.5 µg, i.c.v.). Thirty minutes after pretreatment, ICH was induced by intrastriatal injection of collagenase (0.04 U). Forty-eight hours after ICH, the rats were assessed using a variety of neurobehavioural tests. Subsequently, rats were sacrificed and brains were subjected to gross morphological examination of the haematoma area and histological examination of the damaged area.

Results: Severe neurobehavioural deficits and haematoma with diffuse oedema were observed after intrastriatal collagenase injection. Pretreatment with trans-resveratrol partially restored general locomotor activity, muscle strength and coordination, which was accompanied with reduction of haematoma volume by 73.22% (P < 0.05) and damaged area by 60.77% (P < 0.05) in comparison to the vehicle-pretreated ICH group. The trans-resveratrol-induced improvement in neurobehavioural outcomes and morphological features of brain tissues was inhibited by DPCPX pretreatment.

Conclusion: This study demonstrates that the A1R activation is possibly the mechanism underlying the trans-resveratrol-induced neurological and neurobehavioural protection in rats with ICH.