Melatonin Supplementation During In Vitro Maturation and Development Supports the Development of Porcine Embryos.

Melatonin has been reported to improve the in vitro development of embryos in some species. This study was conducted to investigate the effect of melatonin supplementation during in vitro maturation (IVM) and development culture on the development and quality of porcine embryos. In the first experiment, when the in vitro fertilized embryos were cultured with different concentrations of melatonin (0, 10, 25 and 50 ng/ml) for 8 days, the blastocyst formation rate of embryos cultured with 25 ng/ml melatonin (10.7%) was significantly increased (p < 0.05) compared to the control embryos cultured without melatonin (4.2%). The proportion of DNA-fragmented nuclei in blastocysts derived from embryos cultured with 50 ng/ml melatonin was significantly lower (p < 0.05) than that of embryos cultured without melatonin (2.1% vs 7.2%). In the second experiment, when oocytes were cultured in the maturation medium supplemented with different concentrations of melatonin (0, 10, 25 and 50 ng/ml), fertilized and then cultured with 25 ng/ml melatonin for 8 days, there were no significant differences in the rates of cleavage and blastocyst formation among the groups. However, the proportions (2.7-5.4%) of DNA-fragmented nuclei in blastocysts derived from oocytes matured with melatonin were significantly decreased (p < 0.05) compared to those (8.9%) from oocytes matured without melatonin, irrespective of the concentration of melatonin. Our results suggest that supplementation of the culture media with melatonin (25 ng/ml) during IVM and development has beneficial effects on the developmental competence and quality of porcine embryos.

The combination of hyperthermia or chemotherapy with gimeracil for effective radiosensitization.

PURPOSE: 5-chloro-2,4-dihydroxypyridine (gimeracil) is a component of the oral fluoropyrimidine derivative S-1. Gimeracil was originally added to S-1 to yield prolonged 5-fluorouracil (5-FU) concentrations in serum and tumor tissues by inhibiting dihydropyrimidine dehydrogenase, which degrades 5-FU. We previously demonstrated that gimeracil enhances the efficacy of radiotherapy through the suppression of homologous recombination (HR) in DNA double strand repair. The goal of this paper was to examine the effects of gimeracil on the sensitivity of anticancer drugs and hyperthermia in order to obtain effective radiosensitization. MATERIALS AND METHODS: Various cell lines, including DLD 1 (human colon carcinoma cells) and cells deficient in HR or nonhomologous end-joining (NHEJ), were used in clonogenic assays. The survival of these cells after various treatments (e.g., drug treatment, heat treatment, and radiation) was determined based on their colony-forming ability. RESULTS: Gimeracil enhanced cell-killing effects of camptothecin (CPT), 5-FU, and hydroxyurea. Gimeracil sensitized effects of CPT or 5-FU to cells deficient in HR or NHEJ to a similar extent as in other cells (DLD1 and a parent cell), indicating that its sensitizing mechanisms may be different from inhibition of HR or NHEJ. Combination of gimeracil and CPT or 5-FU sensitized radiation more effectively than each modality alone. Gimeracil also enhanced heat sensitivity at 42°C or more. The degree of heat sensitization with gimeracil increased as the temperature increased, and the combination of gimeracil and heat-sensitized radiation was more effective than each modality alone. CONCLUSION: Gimeracil enhanced sensitivity of CPT, 5-FU, and hyperthermia. Combination of these modalities sensitized radiation more efficiently than each modality alone.

Measurement of urinary zearalenone concentrations for monitoring natural feed contamination in cattle herds: On-farm trials.

The aims of the present study were to investigate the efficacy of measuring bovine urinary zearalenone (ZEN) concentrations by using a commercially available ELISA method in cattle kept under different feeding conditions to monitor the natural contamination of feeds at the farm level, and to investigate the effects of supplementation of a mycotoxin adsorbent (MA) product in the feed based on urinary ZEN concentration. First, Japanese Black cattle herds kept for breeding (4 herds) and fattening (4 herds) purposes were provided with similar feeding conditions. Then, urinary samples from 5 cows in each herd were collected and analyzed. Second, dairy cows from 1 herd fed with total mixed rations (TMR) were selected. After thorough mixing of the MA (40 g/d) with TMR, the supplemented TMR was fed according to the following schedule: with MA for 2 wk, without MA for 3 wk; then with MA for 2 wk and without MA for 6 wk. Urine samples were collected from cows (n = 6 to 7) and examined before and after each interval. Zearalenone concentrations were measured by the ELISA and liquid chromatography-tandem mass spectrometry methods. The concentration of ZEN and its metabolites was expressed after creatinine (Crea) correction [ZEN or metabolites (pg/mL)/Crea (mg/dL); pg/mg of Crea]. In the first experiment, the urinary concentrations of ZEN and its metabolites were variable in all herds, and significant differences were observed between herds. In 1 fattening herd, in particular, urinary ZEN concentrations were greater (P < 0.001) than in the other 3 herds. This might reflect significant natural ZEN contamination of the feed at the farm level. In Exp. 2, urinary ZEN concentrations displayed peculiar trends after supplementation with MA. After 2 wk of supplementation, a significant decrease of ZEN (P < 0.05) was observed. Zearalenone concentrations remained at a reduced amount during 3 wk without MA supplementation and 2 wk with MA supplementation. When MA was not added to the feed for the next 6 wk, the concentrations increased to the original quantity. These findings indicate the usefulness of measuring concentrations of urinary ZEN and its metabolites not only for monitoring the natural ZEN contamination of cattle feed at the farm level but also for in vivo evaluation of MA function after supplementing feeds with MA.

Four novel withanolide-type steroids from the leaves of Solanum cilistum.

Four novel withanolide-type steroids named cilistols p, pm, p1 and u (1-4, respectively), were isolated from the leaves of Solanum cilistum. The respective structures were characterized by spectroscopic means as follows: cilistol p (1) was (22R,24R,25R,26S)-1-oxo-22,26-epoxy-3alpha,5alpha-cycloergostane-6beta,17alpha, 24,25,26-pentaol 26-O-beta-D-glucopyranoside, cilistol pm (2) corresponded to the 6-O-methyl ether derivative of 1; cilistol p1 (3) was represented as the 24-O-methyl ether of 1, and cilistol u (4) was shown to be the epoxide between C-24 and -25, presumably bearing cilistols p, pm and p1 by ring-opening.

Effects of high concentrations of energy sources and metabolites on suspension culture of Chinese hamster ovary cells producing tissue plasminogen activator.

A high-concentration medium was developed for the high-density cultivation of Chinese hamster ovary (CHO) cells to produce tissue plasminogen activator (tPA). First, the effects of the concentrations of several medium components on the growth and metabolism of CHO cells were studied particularly in relation to the serum concentration, in order to develop a suitable concentrated medium. High concentrations of serum, glucose, lactate, and glutamine and a low ammonia concentration increased the specific growth rate and decreased the specific tPA production rate. Among these components, the effect of the serum concentration was the most significant. There was an inverse correlation between the specific rates of cell growth and tPA production. Using a medium containing glutamine at twice the concentration of the original medium (F-12) allowed the medium volume to be reduced to seven-ninths that of the original, without any reduction in tPA productivity. When the F-12 medium was supplemented five-fold with amino acids and vitamins, the total amount of tPA produced, specific rate of tPA production, and yield of tPA per unit volume of medium employed were respectively enhanced 1.7, 1.5, and 3.6 times. As a consequence, the frequency of medium exchange could be reduced. The results strongly indicate that the supplementation of nutrient amino acids and vitamins to a medium with a low concentration of serum are the most critical factors for cost-effective tPA production in high-density cell cultivation.

DNA sensing on a DNA probe-modified electrode using ferrocenylnaphthalene diimide as the electrochemically active ligand.

Naphthalene diimide derivative 1 carrying ferrocenyl moieties at the termini of imide substituents binds intact calf thymus DNA 4 times more strongly than the denatured DNA, and its complex with the intact DNA dissociates 80 times more slowly than that with the denatured DNA. On the basis of these observations, ligand 1 was applied to a probe of electrochemical DNA sensing. A thiol-linked single-stranded DNA probe was immobilized through the S-Au bonding to 20-30 pmol/mm2 on a gold electrode. Following hybridization with the complementary DNA, the electrode was soaked in a solution containing 1 (intercalation step) and then washed with buffer for 5 s. The cyclic voltammogram and differential pulse voltammogram for this electrode gave an electrochemical signal due to the redox reaction of 1 that was bound to the double-stranded DNA on the electrode. Thus, dA20 and the yeast choline transport gene were quantitated at the subpicomole level. The sensitivity of DNA detection was improved to 10 zmol by reducing the amount of immobilized DNA probe and protecting the uncovered surface of the electrode with 2-mercaptoethanol.

[An early-onset case of acute disseminated encephalomyelitis with bilateral thalamic lesions on MRI].

We report the case of 5-year-old girl with acute disseminated encephalomyelitis (ADEM), whose MRI showed bilateral thalamic lesions. She suffered from left optic neuritis and generalized convulsion. Examination of cerebrospinal fluid revealed elevation of mononuclear cells and myelin basic protein (MBP). MRI showed the swelling of left optic nerve and high intensity areas of bilateral thalamus. After methylprednisolone pulse therapy, her visual acuity was dramatically improved and bilateral thalamic lesions were decreased. In childhood, bilateral thalamic lesions were observed in several diseases, such as viral encephalitis. Reye syndrome, Leigh syndrome and acute necrotizing encephalopathy. Demyelinating diseases involving the grey matter were very rare, but we must consider the presence of symmetrical thalamic involvement in patients with ADEM.

Pathologic findings of silent, small hyperintense foci in the basal ganglia and thalamus on MRI.

We analyzed the pathologic features of silent, small hyperintense foci in the basal ganglia and thalamus on MRI T2-weighted images (T2WI). Eight foci were histologically lacunar infarcts (LI), and 21 were dilated perivascular spaces (DPS). The foci with smooth margins were mostly DPS, whereas those with irregular margins were mostly LI (p = 0.018). Most putaminal foci were DPS, whereas most thalamic foci were LI (p = 0.001). The mean areas of LI and DPS overlapped below 19.6 mm2. The shape and site of hyperintense foci on T2WI are important for differentiating LI from DPS.

Expression of stem cell factor (SCF) and SCF receptor (c-kit) in synovial membrane in arthritis: correlation with synovial mast cell hyperplasia and inflammation.

OBJECTIVE: Stem cell factor (SCF), the ligand for the SCF receptor (c-kit) expressed on precursors and mature mast cells (MC), is a major agonist for human MC (e.g., SCF induces MC development, chemotaxis, activation, proliferation of MC precursors, mediates MC adhesion, and changes MC releasability). We investigated expression of SCF and c-kit in synovial membrane with particular reference to the mechanism of local MC hyperplasia and inflammation in arthritis. METHODS: We conducted single and double labeling immunohistochemistry (ABC, APAAP, indirect immunofluorescence techniques) with antibodies to SCF, c-kit, MC tryptase, Ki-67 antigen (marker for proliferating cells), and CD68 (monocyte/macrophage marker). Synovial specimens analyzed were from 31 patients: traumatic arthritis (TrA, n=9), osteoarthritis (OA, n=12), and rheumatoid arthritis (RA, n=10). Control experiments were performed on human lung, skin, and buccal mucosa tissues, on the HMC-1 mast cell line, and isolated lung MC. Morphometry was performed by computerized image analysis. RESULTS: Synovial c-kit expression was found to be restricted to MC, whereas SCF is detected in synovial lining cells, stromal fibroblasts, monocyte/macrophages, endothelial cells, and in vascular basement membranes. SCF staining was localized to MC as well, but it was not possible to specify whether this represents SCF produced by or bound (via c-kit) to MC. In inflamed synovial membranes/areas, SCF was found to be redistributed into the extracellular matrix. Redistribution of SCF was accompanied by degranulation and/or accumulation of c-kit+ MC, the hyperplasia of which correlated positively with histologic inflammation/inflammatory cell densities, but did not appear to involve MC proliferation in situ. These findings appeared to be common for all the conditions (TrA, OA, RA) studied. CONCLUSION: In addition to the demonstration/characterization of SCF and c-kit protein expression in human synovium, results of this study suggest the hypothesis that, in arthritis, local mobilization of SCF may play a role in the development of synovial MC hyperplasia without inducing in situ proliferation of MC, and that the synovial SCF/MC c-kit system may contribute to the local nonspecific inflammatory response/arthritic flares in TrA, OA, and RA.

Vitamin E ameliorates the renal injury of Dahl salt-sensitive rats.

Recently, hyperlipidemia as well as hypertension has been observed in Dahl salt-sensitive (S) rats. In this study, to investigate whether the lipid abnormality is involved in the renal injury of Dahl S rats, we examined the effect of vitamin E on glomerular sclerosis, as vitamin E is an inhibitor of lipid oxidation. Dahl S rats were given a high salt diet (8% NaCl) containing either normal vitamin E (2 mg/100 g) or high vitamin E (50 mg/100 g) for 4 weeks. Dahl salt-resistant (R) rats were given a high salt and normal vitamin E diet. The blood pressure in the Dahl rats increased and was not suppressed by the vitamin E supplement. Serum cholesterol and triglycerides in Dahl S rats were higher than in Dahl R rats at both 0 and 4 weeks. Vitamin E lowered the serum cholesterol level in Dahl S rats at 4 weeks (126 +/- 5 v 150 +/- 12 mg/dL, P < .01). Urinary protein excretion and serum creatinine increased in Dahl S rats, and vitamin E inhibited the increases significantly (urinary protein, 70.7 +/- 0.9 v 178.0 +/- 8.8 mg/day, P < .01; serum creatinine, 0.45 +/- 0.02 v 0.63 +/- 0.05 mg/dL, P < .01). Serum lipid peroxide (LPO) was higher in Dahl S rats than in Dahl R rats, and vitamin E lowered LPO in Dahl S rats (2.10 +/- 0.03 v 2.70 +/- 0.04 nmol/mL, P < .01). In the histologic study, sclerosing score (SS) of glomeruli, which represents the degree of glomerulosclerosis semiquantitatively, was higher in Dahl S rats than in Dahl R rats. Vitamin E lowered SS (114 +/- 3 v 157 +/- 6, P < .01) and ameliorated arterial injuries such as medial thickness with partial necrosis and severe fibrinoid proliferation with inflammatory cell infiltration. In all rats, SS was strongly correlated with urinary protein (r = 0.93, P < .01), serum cholesterol (r = 0.86, P < .01), and serum LPO (r = 0.89, P < .01). These results suggest that the renal injury in Dahl S rats is caused not only by hypertension but also by hyperlipidemia. Therefore, vitamin E might ameliorate the renal damage by inhibiting the oxidation of lipids.

Evaluation of antioxidant activity by chemiluminescence.

A sensitive and simple chemiluminescent method for measuring antioxidant activity was developed. The method is based on antioxidant-dependent quenching of chemiluminescence generated from lipid hydroperoxide and isoluminol/microperoxidase reagent. This method was used to evaluate the antioxidant ability of various antioxidants by measuring the half-inhibition concentration (IC50). The results were compared to those measured with an oxygen electrode method. They were approximately similar in pattern and level, although there was some discrepancy, which was assumed to be due to a possible difference in the reaction mechanisms of the two methods. This method also suggested an additive property of antioxidant activity with different antioxidants. Thus, the present method was used to evaluate the total antioxidant activity in biological tissues such as rat serum, human saliva, and green tea.

Molecular cloning and characterization of human non-smooth muscle calponin.

cDNA clones encoding a calponin isoform with 309 amino acids have been isolated from human heart. The deduced amino acid polypeptide (M(r) 33,697) showed a neutral isoelectric point of 7.1. The mRNA, expressed in cultured smooth muscle cells as well as in fibroblasts, vascular endothelial cells, and keratinocytes, contains a 3' untranslated region of 1.2 kilobases that includes an Alu repetitive sequence in the antisense direction. On the basis of the nucleotide sequence identity to an expressed sequence tag, HUM21ES93 [Cheng, J.-F., Boyartchuk, V., and Zhu, Y. (1994) Genomics 23, 75-84], the human neutral calponin gene is assigned to chromosome 21q11.1. The amino acid sequence indicates that this protein is the human equivalent of mouse calponin-h2 (94.8% identity) [Strasser, P., Gimona, M., Moessler, H., Herzog, M., and Small, J.V. (1993) FEBS Lett. 330, 13-18]. Three tandem repeats of 29 amino acids, a Vav-homologous region and an actin-binding sequence, originally identified in the basic calponin isoform, are conserved. There are two consensus phosphorylation sites for tyrosine kinase. An immunoreactive form of the neutral calponin appears to be localized with vinculin in the cell-to-cell junctions of cardiomyocytes. Mouse calponin-h2 is also expressed in both embryonic and adult heart. These results indicate that the human neutral calponin is a non-smooth muscle isoform, and may play a physiological role in cytoskeletal organization.

[Clinical evaluation of S-1108 in pediatric field].

Nine pediatric patients with bacterial infections (5 cases of tonsillitis, 3 cases of impetigo and 1 case of UTI) were treated with S-1108, and the efficacy and the safety were evaluated. The clinical responses to S-1108 treatment were excellent in 7 cases and good in 2. The efficacy rate was 100%. Bacteriologically, the causative organisms (Streptococcus pyogenes, Staphylococcus aureus, Haemophilus parainfluenzae and Escherichia coli) were eradicated. No clinical side effects were observed. Elevation of CK in 2 cases and eosinophilia in 1 case were noted.

[Environmental and immunological survey of sensitization with Japanese cedar (Cryotomeria japonica) pollen in children. Part 2. The clinical significance of Japanese cedar (Cryptomeria japonica) pollen specific IgE antibody and IgG4 antibody].

The clinical manifestations and Japanese cedar (sugi in Japanese) pollen specific antibodies were studied in 340 children who had not received specific hyposensitization. Specific IgE antibodies were measured by radioallergosorbent test (RAST) and IgG4 antibody by enzyme linked immunosorbent assay (ELISA). There were 52 children (15%) with RAST scores of over 2, and 44 children (13%) with ELISA of over 101 mu/ml. The specific IgG4 level was significantly higher in the RAST positive group than in the negative group. The incidence of pollinosis among children with RAST negative and IgG4 below 100 mu/ml was 13.3%, while combinations of RAST negative and higher ELISA, RAST positive and low ELISA, and RAST positive and higher ELISA were 50.0+-62.5%. There were 5 children with RAST scores of 4, and 6 children with ELISA of over 201 mu/ml. Symptoms of pollinosis manifested in all of them. Since the children with low IgE and high IgG4 antibodies had comparatively more symptoms, the IgG4 antibodies might be reaginic. It was concluded that measurement of sugi specific IgG4 antibodies as well as IgE antibodies could provide useful data on pollinosis in children.

[Clinical significance of serum erythropoietin levels in patients with multiple myeloma].

Serum erythropoietin (Epo) levels were measured in 53 patients with multiple myeloma (MM), 49 normal subjects and 53 patients with some hematological diseases including aplastic anemia (AA), iron deficiency anemia, etc. to study the significance of erythropoietin in anemia of MM. The serum Epo level was 72.0 +/- 94.4 mIU/ml (mean +/- SD) in MM patients, which was significantly higher than in normal subjects (24.1 +/- 6.1 mIU/ml), but lower than in AA patients (7069.9 +/- 9406 mIU/ml). A significant inverse correlation was found between the hemoglobin (Hb) levels and the logarithmic values of serum Epo levels (r = -0.543, p < 0.05) in MM patients. This inverse correlation was stronger (r = -0.636, p < 0.05) in MM patients without renal dysfunction than in whole MM patients, while no correlation was observed in MM patients with renal dysfunction. These results indicate that MM patients with renal dysfunction have a low ability to synthesize Epo and that the supplemental therapy of recombinant Epo is effective to improve their anemia. In addition, the circadian rhythm of serum Epo level was lower in the morning than in the afternoon in both MM patients and normal controls. Serum Epo levels after chemotherapy in MM patients were elevated temporarily and then decreased in spite of no change of blood Hb level.

[The pharmacokinetics of cisplatin and its influence on renal function according to different infusion methods (report II)--alleviation of renal impairment by bismuth subnitrate combined with ginseng and tang-koui ten].

In the previous report, we discussed in vivo dynamics and renal impairment resulting from the difference in cisplatin CDDP administration. In the present study, we investigated in vivo dynamics and renal impairment under 1 h of continuous drip infusion of CDDP. The free-type Pt concentration ratio, which is considered to indicate antitumor activity in the overall plasma Pt concentration at the end of the administration, was noted to be approximately 64%. This was significantly higher than other administration methods, making this an effective administration procedure. However, NAG, which is considered to be an index of renal impairment along with the amount of beta 2-microglobulin urine excretion, showed higher values than those by other administration methods. Thus, using the 1-hr continuous CDDP drip infusion, we attempted to alleviate the renal impairment, combining bismuth subnitrate with ginseng and Tang-koui ten. As a result, the amount of beta 2-microglobulin excretion in the urine was diminished as well as NAG, and a reduction of the renal impairment was noted. These alleviational effects of the renal impairment were attributed to the method of consistent oral administration by which alleviation could take place with little load on the patient.

Ultrastructural demonstration of increased sulfated proteoglycans and calcium associated with chondrocyte cytoplasmic processes and matrix vesicles in rat growth plate cartilage.

A monoclonal antibody (3-B-3) to chondroitin 6-sulfated proteoglycan was used with immunoperoxidase electron microscopy to study the relationship of chondrocyte cytoplasmic processes and matrix vesicles in rat epiphyseal growth plate cartilage. Immunoperoxidase staining of the chondrocyte plasmalemma was found at all levels in the growth plate and was most prominent in the hypertrophic zone. The plasmalemma and matrix of the cytoplasmic process often demonstrated stronger reactivity than the remainder of the cell surface. Matrix vesicles showed weak to strong surface or internal reactivity. The majority of them stained very similarly to the cytoplasmic process. X-ray microanalysis of specimens processed by rapid freezing and freeze substitution confirmed that both sulfur and calcium were localized within or in close association with both the cytoplasmic process and the matrix vesicle, suggesting a chemical combination of calcium with sulfated proteoglycans at both sites. These results indicate that there is a selective increase in the concentration of membrane-associated sulfated proteoglycan and calcium in the cell process, from which matrix vesicles may be released into the extracellular matrix.

Time course analyses of kinins and other mediators in plasma exudation of rat kaolin-induced pleurisy.

Pleurisy was induced in rats by an intrapleural injection of 0.5 ml of 1% kaolin. The exudation of plasma into the pleural cavity showed two peaks at 20 min and 3-5 h after the kaolin injection. The volume of the pleural fluid increased gradually up to 5 h. The effects of treatment with mepyramine, methysergide, captopril, bromelain and indomethacin suggested that the early phase (20 min) of exudation was mediated mainly by kinins, histamine and 5-HT, and that the late phase (3 h) was mediated by prostaglandins (PGs) and possibly kinins. We measured the levels of histamine, kinin and PG in the pleural exudate to verify the involvement of the mediators mentioned above. Intracellular histamine levels decreased markedly and extracellular histamine levels increased significantly 20 min after the induction of kaolin pleurisy. Only threshold levels of kinin were detected after the induction of pleurisy. Captopril treatment, however, increased kinin levels which peaked at 20 min and decreased rapidly thereafter. Levels of 6-keto-PGF1 alpha and thromboxane B2 showed a peak at 20 min, whereas levels of PGE2 increased gradually from 20 min to 5 h. These results indicate that kaolin-induced pleurisy is a kinin-related inflammation and could be used as a model for studying the in vivo interaction of the kallikrein-kinin system and PGs at inflammatory sites.

SEM-EPMA observation of three types of apatite-containing glass-ceramics implanted in bone: the variance of a Ca-P-rich layer.

The progressive changes of a Ca-P-rich layer between bone and three types of apatite-containing glass-ceramics of the same chemical composition: MgO 4.6, CaO 44.9, SiO2 34.2, P2O5 16.3, CaF2 0.5 (in weight ratio) were examined. Plates (15 mm X 10 mm X 2 mm, mirror surface) containing apatite (35 wt%) (designated A-GC), apatite (35 wt%) and wollastonite (40 wt%) (designated A.W-GC), and apatite (20 wt%), wollastonite (55 wt%), and whitlockite (15 wt%) (designated A.W.CP-GC) were prepared. They were implanted into the tibia of mature male rabbits for 5 days, 10 days, 20 days, 30 days, 60 days, 6 months, and 12 months. All three types of glass-ceramics showed direct bonding to the bone 30 days after implantation. It was observed by SEM-EPMA 30 days after implantation that Si and Mg content decreased, P content increased, and Ca content did not change across the reactive zone from the glass-ceramics to bone. The level of P and Si in the A.W.CP-GC changed five days after implantation. In A.W-GC and A-GC, a little change in P and Si levels was observed between 10 and 20 days after implantation. The width of reactive zone was narrowest with A-GC, wider with A.W-GC, and widest with A.W.CP-GC. The dissolution of glass-ceramics stopped 6 months after implantation. This phenomenon shows that the glass-ceramics may be suitable for clinical use.