Anti-Inflammatory Activities of Inotilone from Phellinus linteus through the Inhibition of MMP-9, NF-κB, and MAPK Activation In Vitro and In Vivo

Inotilone was isolated from Phellinus linteus. The anti-inflammatory effects of inotilone were studied by using lipopolysaccharide (LPS)-stimulated mouse macrophage RAW264.7 cells and λ-carrageenan (Carr)-induced hind mouse paw edema model. Inotilone was tested for its ability to reduce nitric oxide (NO) production, and the inducible nitric oxide synthase (iNOS) expression. Inotilone was tested in the inhibitor of mitogen-activated protein kinase (MAPK) [extracellular signal-regulated protein kinase (ERK), c-Jun NH2-terminal kinase (JNK), p38], and nuclear factor-κB (NF-κB), matrix-metalloproteinase (MMP)-9 protein expressions in LPS-stimulated RAW264.7 cells. When RAW264.7 macrophages were treated with inotilone together with LPS, a significant concentration-dependent inhibition of NO production was detected. Western blotting revealed that inotilone blocked the protein expression of iNOS, NF-κB, and MMP-9 in LPS-stimulated RAW264.7 macrophages, significantly. Inotilone also inhibited LPS-induced ERK, JNK, and p38 phosphorylation. In in vivo tests, inotilone decreased the paw edema at the 4th and the 5th h after Carr administration, and it increased the activities of catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPx). We also demonstrated that inotilone significantly attenuated the malondialdehyde (MDA) level in the edema paw at the 5th h after Carr injection. Inotilone decreased the NO and tumor necrosis factor (TNF-α) levels on serum at the 5th h after Carr injection. Western blotting revealed that inotilone decreased Carr-induced iNOS, cyclooxygenase-2 (COX-2), NF-κB, and MMP-9 expressions at the 5th h in the edema paw. An intraperitoneal (i.p.) injection treatment with inotilone diminished neutrophil infiltration into sites of inflammation, as did indomethacin (Indo). The anti-inflammatory activities of inotilone might be related to decrease the levels of MDA, iNOS, COX-2, NF-κB, and MMP-9 and increase the activities of CAT, SOD, and GPx in the paw edema through the suppression of TNF-α and NO. This study presents the potential utilization of inotilone, as a lead for the development of anti-inflammatory drugs

Biochemical and Histopathological Changes in the Mortality Caused by Acute Ischemic Limb Injury: A Rabbits' Model

Restoration of blood flow to an acute ischemic extremity may deteriorate the ischemic injury, lead to multiple organ dysfunction or even death. This paradox of continuing injury during reperfusion is not completely understood. The role of multi-organ damage in the mortality caused by ischemic limb injury is also still not clarified. The purpose of this study is to determine the biochemical and histopathological changes in the mortality caused by ischemic limb injury. After anesthesia, the hindlimbs of 14 New Zealand white rabbits were made ischemic and set into 8 hours or 12 hours of ischemia. Blood samples were obtained then the creatine kinase (CK) levels were determined and CK isoenzymes analyzed. All rabbits with 8 hours' ischemia survived well, and 5 of the 7 rabbits with 12 hours' ischemia expired within 8 hours after reperfusion. CK elevation was correlated most strongly with the time of the ischemic insults. The percentage of CK-MB isoenzyme remained unchanged after 8 hours' ischemia- reperfusion insult, while increased significantly after 12 hours' ischemia -reperfusion insult. Histologic examinations showed that the major systemic manifestation was massive destruction of the liver and kidney. The injuries are more obvious in areas with the greatest blood flow during reperfusion. We concluded that the ratio of CK-MB isoenzyme is most useful for distinguishing the risk of mortality caused by acute ischemic limb injury, and the cause of systemic complications are attributed to the multi-organ failure

Chromaffinity, uranaffinity and argentaffinity of small granule-containing (SGC) cells in rat superior cervical ganglia

A systemic examination on the small granule-containing (SGC) cells in rat superior cervical ganglia was conducted by conventional and cytochemical electron microscopy including chromaffin, argentaffin and uranaffin reactions. According to the fine structure of dense cored vesicles (DCVs) in the cytoplasm, three types of small granule-containing (SGC) cells were revealed - Type 1: 90 - 160 nm vesicles with cores of moderate or low electron density; Type 11: 130 - 330 nm vesicles, polymorphic with highly electron dense cores; Type 111: elongated vesicles (170 nm x 60 nm) with cores of moderate to low electron density. The majority of SGC cells were the Type 1 cells (78%) and Type 11 and 111 cells made up 13% and 9% of SGC cell population, respectively. Cytochemical results demonstrated that only the Type 11 cells displayed a positive chromaffin reaction and al1 three types of SGC cells showed argentaffinity and uranaffinity. The present study is the first to demonstrate the argentaffin reaction at ultrastructural level in SGC cells of sympathetic ganglia. Based on the results of the present study we also concluded that (1) the DCVs of Type 11 SGC cells contained noradrenaline and (2) biogenic amines and nucleotides (ATPs) coexisted in the DCVs of al1 three types of SGC cells

The Expression of α-Internexin and Peripherin in the Developing Mouse Pineal Gland

The mammalian pineal gland contains pinealocytes, interstitial glial cells, perivascular macrophages, neurons and neuron-like cells. The neuronal identity of neurons and neuron-like cells was an enigma. α- Internexin and peripherin are specific neuronal intermediate filament proteins and are expressed differentially in the CNS and PNS . We investigated the development of immunoreactivity and expression patterns of mRNAs for α-internexin and peripherin in the mouse pineal gland to determine the neuronal identity of these cells. Both α-internexin- and peripherin-immunoreactive cells were readily visualized only after birth. Both proteins were at the highest level on the postnatal day 7 (P7), rapidly declined at P14, and obtained their adult level at P21. Both protein and mRNA of α- internexin are expressed in some cells and nerve processes, but not all, of adult mouse pineal gland. Less number of peripherin immunoreactive or RNA-expressing cells and nerve processes were identified. Accumulations of α-internexin and peripherin proteins were also found in the cells from the aged pineal gland (P360). We concluded that some cells in the developing mouse pineal gland may differentiated into neurons and neuron-like cells expressing both α-internexin and/or peripherin only postnatally, and these cells possess dual properties of CNS and PNS neurons in nature. We suggested that they may act as interneurons between the pinealocyte and the distal neurons innervating the pinealocytes, or form a local circuitry with pinealocytes to play a role of paracrine regulatory function on the pinealocytes

Effects of Zinc Deficiency on the Vallate Papillae and Taste Buds in Rats

BACKGROUND AND PURPOSE: Zinc deficiency is associated with multiple clinical complications, including taste disturbance, anorexia, growth retardation, skin changes, and hypogonadism. We investigated the zinc-deficiency-induced morphologic changes in the vallate taste buds of weanling and young adult male Wistar rats. METHODS: A total of 24 weanling and 30 young adult rats were used. Each age group was further divided into a control group fed a zinc-adequate (50 ppm) diet, a zinc-deficient (< 1 ppm) diet group, and a zinc-adequate pair-fed group who were fed the same amount of food as that taken by the zinc-deficient group. Weanling rats were fed for 4 weeks and young adult rats were fed for 6 weeks. The morphometry and morphologic changes of vallate taste buds were analyzed using light and transmission electron microscopy. RESULTS: Light microscopy revealed no significant difference in papilla size and morphology among the various groups. In both weanling and young adult rats in the zinc-deficient diet and pair-fed groups, the number of taste buds per papilla (per animal) and the average profile area of the taste bud were significantly smaller than those of the corresponding controls (p < 0.05). Ultrastructural changes were seen only in the taste buds of weanling rats fed the zinc-deficient diet, with derangement of the architecture of the taste bud and widening of the intercellular space between taste bud cells. The proportion of type I taste bud cells in the taste buds of weanling rats fed the zinc-deficient diet decreased from 59% to 39%, and that of type II taste bud cells decreased from 25% to 12%. No obvious changes in the ultrastructure of type III taste bud cells were observed. CONCLUSIONS: The main effects of zinc deficiency in weanling and young adult rats and in adequate diet pair-fed rats were changes in the number and size of taste buds, and fine structure changes in the taste bud cells, especially during the accelerated growth stage after weaning