Aged garlic extract therapy for sickle cell anemia patients

Background  

Sickle cell anemia is one of the most prevalent hereditary disorders with prominent morbidity and mortality. With this disorder oxidative, phenomena play a significant role in its pathophysiology. One of the garlic (Allium sativum L.) formulations, aged garlic extract (AGE), has been reported to exert an anti-oxidant effect in vitro, we have evaluated the anti-oxidant effect of AGE on sickle red blood cells (RBC).     

Dietary combination of sucrose and linoleic acid causes skeletal muscle metabolic abnormalities in Zucker fatty rats through specific modification of fatty acid composition

A dietary combination of sucrose and linoleic acid strongly contributes to the development of metabolic disorders in Zucker fatty rats. However, the underlying mechanisms of the metabolic disorders are poorly understood. We hypothesized that the metabolic disorders were triggered at a stage earlier than the 8 weeks we had previously reported. In this study, we investigated early molecular events induced by the sucrose and linoleic acid diet in Zucker fatty rats by comparison with other combinations of carbohydrate (sucrose or palatinose) and fat (linoleic acid or oleic acid). Skeletal muscle arachidonic acid levels were significantly increased in the sucrose and linoleic acid group compared to the other dietary groups at 4 weeks, while there were no obvious differences in the metabolic phenotype between the groups. Expression of genes related to arachidonic acid synthesis was induced in skeletal muscle but not in liver and adipose tissue in sucrose and linoleic acid group rats. In addition, the sucrose and linoleic acid group exhibited a rapid induction in endoplasmic reticulum stress and abnormal lipid metabolism in skeletal muscle. We concluded that the dietary combination of sucrose and linoleic acid primarily induces metabolic disorders in skeletal muscle through increases in arachidonic acid and endoplasmic reticulum stress, in advance of systemic metabolic disorders.     

Elevated urinary plasmin activity resistant to alpha2-antiplasmin in acute poststreptococcal glomerulonephritis.

BACKGROUND: A pathogenic role of intraglomerular plasmin bound to nephritogenic antigen (nephritis-associated plasmin receptor, NAPlr) and resistant to physiologic inhibitors such as alpha(2)-antiplasmin (alpha(2)-AP) has recently been proposed in acute poststreptococcal glomerulonephritis (APSGN). To confirm this concept, we analysed the urinary profile of plasmin cascade in APSGN patients. METHODS: Urine samples from 10 patients with APSGN, 12 patients with IgA nephropathy (IgAN), 10 patients with streptococcal infection without nephritis (SI) and 10 healthy control subjects were analysed. The alpha(2)-AP-resistant plasmin activity was assessed by a chromogenic assay after alpha(2)-AP was added to each urine sample. Urinary plasminogen activator (PA) and plasmin were further analysed by polyacrylamide gel zymography. Urinary NAPlr was assessed by western blot analysis in selected samples. RESULTS: Urinary alpha(2)-AP-resistant plasmin activity corrected for creatinine concentration (units/g x creatinine) was significantly higher in patients with APSGN (2.99 +/- 0.63) than in patients with IgAN (1.02 +/- 0.20, P < 0.01), SI (0.79 +/- 0.17, P < 0.01), or in healthy control subjects (0.73 +/- 0.18, P < 0.01). This tendency was confirmed by casein gel zymography. However urinary PA activity assessed by plasminogen-casein gel zymography did not differ between groups. NAPlr was detected in the urine of APSGN patients. CONCLUSIONS: We found elevated urinary plasmin activity resistant to alpha(2)-AP, which may be due to urinary excretion of NAPlr in patients with APSGN. This result supports the pathogenic role of the NAPlr-plasmin complex in the development of APSGN. Furthermore, alpha(2)-AP-resistant urinary plasmin activity may be useful as a diagnostic marker for APSGN.     

Epidermal Growth Factor Prolongs Survival Time of Tumor-bearing Mice

We observed that human epidermal growth factor (hEGF) alone prolonged the survival time of mice bearing various murine syngeneic tumors as well as athymic nude mice bearing human xenografts. No changes in the subcutaneous solid tumor mass volume were observed. Prolongation of survival time by hEGF was observed in mice bearing murine epidermoid carcinoma (BSC) and human gastric carcinoma (KATO III), but not in murine epidermoid carcinoma (KLN205) or human epidermoid carcinoma (A431). Human tumor cells such as A431, KATO III, and murine tumor cells, KLN205, BSC had roughly 2 × 10⁶, 3 × 10⁴, 1.3 × 10³ and 1 × 10³ EGF receptors/cell, respectively. Although KLN205 and BSC tumor cells maintained nearly the same number of EGF receptors, the effects of hEGF were very different. Although A431 tumor cells had nearly 100 times more receptors than KATO III cells, the prolongation of survival time of mice bearing A431 by hEGF was no better than that of mice bearing KATO III. Accordingly, it appears that this prolongation of survival time by hEGF is independent of the number of EGF receptors on tumor cells. In addition, hEGF was shown to inhibit experimental pulmonary metastasis of murine BSC tumor, but was ineffective with murine KLN205 tumor. These results suggest that prolongation of survival time by hEGF may result from the inhibition of tumor cell metastasis and EGF may play a role in preventing the metastasis of certain malignant neoplasms unrelated to its effects through the EGF receptor on tumor cells.     

Electron microscopic study on regression of cardiac hypertrophy in spontaneously hypertensive rats

In this electron microscopic study of cardiocytes during regression of left ventricular hypertrophy (LVH) in spontaneously hypertensive rats (SHR), nifedipine (SHR-N) and lisinopril (SHR-L) were given to 15-week-old SHR (SHR-C₁₅) for 20 weeks. In untreated SHR, cardiocytes were enlarged, lateral branches (LBs) multiplied and intracellular organelles became degenerated between 15 and 35 weeks of age. In SHR-N, despite a slight reduction in blood pressure (BP), LVH regressed and cell width and the number of LBs were preserved at the level found in SHR-C₁₅. In SHR-L, LVH regressed with moderate suppression of BP; cell width, cell length and the number of LBs reverted to those in agematched Wistar Kyoto rats. In both SHR-N and SHR-L, intracellular ultrastructures were nearly normalized. These findings suggest that, in addition to BP reduction, other pharmacological factors play a role in structural repair of hypertrophied cardiocytes.     

Ultrastructure of a coronary chemoreceptor of dogs and cats

Ultrastructure of a coronary chemoreceptor which causes a hypertensive reflex was examined in three normal dogs and three normal cats. The tissue is a periarterial glomoid body composed of highly vascular, grape-like clusters regularly located in the immediate vicinity of the left main coronary artery, from which it receives its blood supply. Its general histological organization resembles that of the carotid body. Morphometric analysis indicates that chief cells were about twice as numerous as sustentacular cells. The chief cells contain abundant osmiophilic dense granules and are surrounded by the processes of sustentacular cells. These dense granules resemble those of the carotid body and adrenal medulla, suggesting that they also may be involved in the production and storage of amines.Abundant nerve endings and formed synapses are associated with the chief cells. Numerous sympathetic and parasympathetic efferent nerve endings had characteristic presynaptic junctions to chief cells. In contrast the sensory afferents packed with mitochondria but with few synaptic vesicles had characteristic postsynaptic junctions to the chief cells. These findings suggest that the chief cells not only stores amines but also serves as a sensory or chemoreceptive site.     

Age differences in cyclin-dependent kinase inhibitor expression and rb hyperphosphorylation in human corneal endothelial cells

PURPOSE: Human corneal endothelial cells (HCECs) are considered to be nonreplicative in vivo; however, isolated HCECs can be cultured and grown successfully, indicating that they retain proliferative capacity. This capacity to replicate tends to decrease with donor age. Cyclin-dependent kinase inhibitors (CKIs) are important negative regulators of the cell cycle. Of those CKIs, p16INK4a, p21WAF1/Cip1, and p27Kip1 are expressed in corneal endothelium. To help reveal the mechanism of this age-related difference, the relative expression of those CKIs and the kinetics of hyperphosphorylation of the retinoblastoma protein, Rb, were analyzed in HCECs from various aged donors. METHODS: Fresh-frozen sections of corneas from an 18-year-old and a 74-year-old donor were immunostained to reveal the expression and localization of the three CKIs in corneal endothelium in situ. HCECs from eight donors of various ages were isolated and cultured until they reached passage 4. After the cells reached confluence, total protein was extracted, and the relative expression of p16(INK4a), p21WAF1/Cip1, and p27Kip1 was determined by Western blot analysis. A parallel analysis was performed with primary cultures of HCECs obtained from eight different donors. Subconfluent passage 2 HCECs from eight donors were serum starved and, at different times after growth factor stimulation, protein was extracted, and Western blot analysis was used to compare the overall expression of Rb protein and the kinetics of Rb hyperphosphorylation. RESULTS: Immunocytochemistry confirmed the expression and nuclear localization of p16(INK4a), p21WAF1/Cip1, and p27Kip1 in HCECs in situ. Western blot studies revealed an age-related increase in p16INK4a and p21WAF1/Cip1 protein expression in cultured HCECs. Expression of p27Kip1 tended to decrease with the donor's age in passage-4 cells; however, there was no significant difference in p27Kip1 expression level between young and older donors in primary cultured HCECs. No age-related difference in total Rb protein was observed in the Western blots; however, the rate of Rb hyperphosphorylation was significantly slower in HCECs from older donors. CONCLUSIONS: p16(INK4a), p21WAF1/Cip1, p27Kip1, and Rb were all expressed in HCECs, regardless of donor age. Age-related differences in the relative expression of p16INK4a and p21WAF1/Cip1 and in the kinetics of Rb hyperphosphorylation led to the conclusion that, in addition to the normal inhibitory activity of p27Kip1, there is an age-dependent increase in negative regulation of the cell cycle by p16INK4a and p21WAF1/Cip1. This additional molecular mechanism may be responsible, at least in part, for the reduced proliferative response observed in HCECs from older donors.     

Epidermal growth factor receptor-mediated selective cytotoxicity of antitumor agents toward human xenografts and murine syngeneic solid tumors

Severe toxic side effects of antiproliferative agents limit their clinical usefulness as antitumor drugs. Recently we observed that the antitumor efficacy of various antitumor agents (5-fluorouracil, tegafur, adriamycin, mitomycin C, cyclophosphamide, and cisplatin) against experimental solid tumors was enhanced by prior or simultaneous administration of human epidermal growth factor (EGF). However, coadministration of EGF did not enhance the toxicity of antitumor agents as measured by LD50 and body weight loss. The above selective potentiation of efficacy of the antitumor agents by human EGF can be characterized as follows. In a dose-dependent manner, human EGF enhanced the efficacy of an antitumor agent (5-FU) treatment against human epidermoid carcinoma A431 transplanted sc in athymic nude mice [ED50 = 2.9 (0.2-49.7, 95% confidence interval) microgram/kg, sc]. Various degrees of enhancement were also observed against other experimental tumors transplanted sc. The degrees of enhancement were directly proportional to the numbers of human EGF binding sites present on tumor cell plasma membrane (threshold of binding site density = 1.5 X 10(3) sites/cell) using 5-FU or cisplatin as an antitumor agent, thus suggesting that the binding of EGF to the receptors on tumor cells is an essential process in enhancing the susceptibility of tumor cells to antitumor agents. Normal cells including intestinal epithelial and bone marrow cells are endowed with fewer EGF binding sites (less than 10(3) sites/cell). This may explain partially the absence of EGF-enhanced cytotoxicity by antitumor agents toward normal cells.     

A novel gastric lesion model induced in rats by partial gastric vascular ligation

Previous studies have suggested that histamine treatment after gastric vascular ligation induces mucosal damage in the rat stomach. Although ligation of left gastric artery and vein (L-AV) alone did not cause any damage in the stomach within 4 h, but provoked mild lesions due to ischemia 24 h later. In the present study we demonstrated a new model of gastric lesions induced by L-AV ligation and examined the effects of various anti-ulcer drugs on this lesion model. The gastric lesions induced by L-AV ligation occurred at the corpus and antrum, especially at the corpus-antrum border, when examined 24 h later, and the severity of damage reached maximum 3 days after L-AV ligation. Repeated treatment with omeprazole or sucralfate for 3 days significantly prevented the development of gastric lesions induced by L-AV ligation, in whole mucosa, including the antrum. By contrast, famotidine given for 3 days showed a significant protection against total lesions in the whole mucosa, but had no effect on the antral lesions. Both omeprazole and famotidine dose-dependently decreased gastric acid output while sucralfate raised the intraluminal pH due to the acid-neutralizing action. These results suggest that the pathogenesis of gastric lesions induced by L-AV ligation differs depending on the region, the corpus and the antrum, and the lesions occurred in the latter area seem to be resistant to acid suppression. It is assumed that this new model of gastric lesions is useful for screening the drugs that affect gastric mucosal defense rather than acid secretion.