Suppressive effects of 1,4-dihydroxy-2-naphthoic acid administration on bone resorption.

SUMMARY: The main component of the metabolic by-products of fermentation by Propionibacterium freudenreichii ET-3 is 1,4-dihydroxy-2-naphthoic acid (DHNA), which has a naphthoquinone skeleton, as in vitamin K2. This study showed that DHNA improved bone mass reduction with osteoporosis model mice caused by FK506. INTRODUCTION: Growth of the intestinal bacterium Lactobacillus bifidus is specifically facilitated by DHNA. The present study used osteoporosis model mice to investigate the effects of DHNA on bone remodeling. METHODS: FK506, an immunosuppressant, was used to prepare osteoporosis model mice. Thirty mice were divided into three groups: FK group, FK+DHNA group, and control group. In the FK group, FK506 was administered to induce bone mass reduction. In the FK-DHNA group, FK506 and DHNA were administered concurrently to observe improvements in bone mass reduction. To ascertain systemic and local effects of DHNA, we investigated systemic pathological changes in colon, kidney function and cytokine dynamics, and morphological and organic changes in bone and osteoclast dynamics as assessed by culture experiments. RESULTS: Compared to the FK group without DHNA, colon damage and kidney dysfunction were milder for FK+DHNA group, and production of inflammatory cytokines (interleukin (IL)-1beta, IL-6 and tumor necrosis factor (TNF)-alpha) was more suppressed. Furthermore, compared to the group without DHNA, histological analyses and radiography showed that bone resorption was suppressed for the DHNA group. Culture experiments using osteoclasts from murine bone marrow showed osteoclast suppression for the DHNA group compared to the group without DHNA. CONCLUSION: These results show that DHNA has some effects for improving bone mass reduction caused by FK506.

Discriminative disulfide-bonding affinity labeling of opioid receptor subtypes.

The affinity-labeling technique is an extremely important method in receptor biochemistry. The 3-nitro-2-pyridinesulfenyl (Npys) group, attached to a mercapto group, can react only with a free thiol group (the beta-mercapto group of cysteine residue) of the target receptor molecules, forming a disulfide bond. This disulfide bonding is mediated through the thiol-disulfide exchange reaction. Unlike other labeling methods, the approach utilizing such chemically activated thiol-containing ligands is able to reproduce an unlabeled protein by treatment with dithiothreitol, a reducing reagent. This provides several unique aspects for the studies elucidating the structure-function relationships between the peptide and the receptor. Based on the SNpys affinity technique, we have achieved the discriminative disulfide-bonding affinity labeling of the three different subtypes of opioid receptors: mu, delta and kappa. This article reviews our novel affinity techniques in the in vitro receptor biochemistry.

Effect of the reducing agent on the oxygen-inhibited layer of the cross-linked reline material.

The purpose of this study was to clarify the effect of the reducing agent on the oxygen-inhibited layer of the cross-linked reline material. A commercial autopolymerizing reline resin containing 1,6-hexanediol dimethacrylate as cross-linking agent and 1 wt.% sodium sulphite solution as a reducing agent was prepared. The inhibited layer was observed using an optical transmission microscope under the conditions of the application of sodium sulphite for 0, 1, 5 and 15 min after curing for 10 min in air. As a control, the reline material was cured on sealing from air. Moreover, the three-point flexural strength test was performed under the same conditions. The fracture was then observed using scanning electron microscopy (SEM). Although hardness of the inhibited layer was enhanced after the application of the reducing agent, the layer was still observed. The flexural strength of the control and the groups after application of the reducing agent was significantly higher than the group without reducing agent. SEM examination revealed many polymer beads on the group without reducing agent, whereas polymer beads could not be observed on the groups applying the reducing agent. These results indicated that the application of sodium sulphite was effective in hardening the surface unpolymerized zone.

Exploration of universal cysteines in the binding sites of three opioid receptor subtypes by disulfide-bonding affinity labeling with chemically activated thiol-containing dynorphin A analogs.

A ligand containing an SNpys group, i.e. 3-nitro-2-pyridinesulfenyl linked to a mercapto (or thiol) group, can bind covalently to a free mercapto group to form a disulfide bond via the thiol-disulfide exchange reaction. This SNpys chemistry has been successfully applied to the discriminative affinity labeling of mu and delta opioid receptors with SNpys-containing enkephalins [Yasunaga, T. et al. (1996) J. Biochem. 120, 459-465]. In order to explore the mercapto groups conserved at or near the ligand binding sites of three opioid receptor subtypes, we synthesized two Cys(Npys)-containing analogs of dynorphin A, namely, [D-Ala2, Cys(Npys)8]dynorphin A-(1-9) amide (1) and [D-Ala2, Cys(Npys)12]dynorphin A-(1-13) amide (2). When rat (mu and delta) or guinea pig (kappa) brain membranes were incubated with these Cys(Npys)-containing dynorphin A analogs and then assayed for inhibition of the binding of DAGO (mu), deltorphin II (delta), and U-69593 (kappa), the number of receptors decreased sharply, depending upon the concentrations of these Cys(Npys)-containing dynorphin A analogs. It was found that dynorphin A analogs 1 and 2 effectively label mu receptors (EC50 = 27-33 nM), but also label delta receptors fairly well (160-180 nM). However, for kappa receptors they showed drastically different potencies as to affinity labeling; i.e., EC50 = 210 nM for analog 1, but 10,000 nM for analog 2. Analog 2 labeled kappa receptors about 50 times more weakly than analog 1. These results suggested that dynorphin A analog 1 labels the Cys residues conserved in mu, delta, and kappa receptors, whereas analog 2 only labels the Cys residues conserved in mu and delta receptors.

Early site of lymphatic involvement from right renal cell carcinoma: CT demonstration and method of lymphadenectomy.

The value of lymphadenectomy in the treatment of renal cell carcinoma remains controversial. The precise location of nodal metastasis and method of lymphadenectomy had seldom been mentioned. We report 2 patients with right renal cell carcinoma who had regional lymph node involvement without blood-borne metastasis. The involved lymph nodes were demonstrated by CT scan just behind the inferior vena cava at the level of renal hilus. These nodes seemed to be the first sites of nodal involvement from the right renal cell carcinoma. Elevation of the inferior vena cava by dividing its small branches, including a few lumbar veins, was necessary for complete removal of these nodes en bloc with the right kidney.

Ex vivo comparison of radiological and histological evaluation of early metastatic lesions of pelvic lymph nodes from carcinoma of the bladder or prostate.

To study the limitations of lymphangiography in the detection of early lymphatic spread of pelvic malignancies, we evaluated 587 lymph nodes from 23 patients with stages pN0 to 2 carcinoma of the bladder or prostate. Pelvic lymphadenectomy was performed 5 to 10 days after bipedal lymphangiography. Excised lymph nodes were separated one by one and an x-ray of each node was taken (lymphnodegram). The individual lymphnodegram was compared to the histological findings. Interpretations of lymphnodegrams from all 17 nodes with metastases were positive in 5, suspicious in 1, negative in 9 and radiolucent in 2. False negative judgments occurred chiefly because metastatic foci were microscopic. Two lymph nodes without metastasis were interpreted as positive for disease because of fat replacement of the nodes. These results indicated that lymphangiography is not suitable for the detection of early lymphatic metastases of carcinoma of the bladder or prostate.

Right adrenal pheochromocytoma with anterolateral displacement of the inferior vena cava: skin incision and approach.

We report a case of a right adrenal pheochromocytoma with prominent anterolateral displacement of the inferior vena cava. A right ipsilateral anterior subcostal incision extending from the xiphoid process to the 11th intercostal space allowed for wide upward retraction of the right costal arch. This incision is useful for easy mobilization of the liver and manipulation of the inferior vena cava without thoracotomy.