Intracellular GSH of Streptococcus thermophilus shows anti-oxidative activity against low-density lipoprotein oxidation in vitro and in a hyperlipidaemic hamster model.

Streptococcus thermophilus YIT 2001 (ST-1), a lactic acid bacterial strain, was shown to have inhibitory effects on the oxidation of low-density lipoprotein (LDL) and the development of aortic fatty lesions in an animal model, and lower the serum levels of malondialdehyde-modified LDL, an oxidative modification product of LDL, in a clinical trial. This study aimed to identify the intracellular active component of ST-1 associated with anti-oxidative activity against LDL oxidation. High-performance liquid chromatography-electrospray ionisation mass spectrometry analysis after fractionation of the cellular extract by reversed-phase chromatography demonstrated that the active fraction contained reduced glutathione (GSH). GSH showed anti-oxidative activity in a dose-dependent manner, while this activity disappeared following thiol derivatisation. ST-1 had the strongest anti-oxidative activity against LDL oxidation and the highest level of intracellular GSH among five strains of S. thermophilus. In addition, the anti-oxidative activity of ST-1 after thiol derivatisation decreased by about half, which was similar to that of three other strains containing poor or no intracellular GSH or thiol components. Moreover, anti-oxidative activity against LDL oxidation was observed in hyperlipidaemic hamsters fed with high GSH ST-1 cells but not in those given low GSH cells. These findings suggest that intracellular GSH in ST-1 may provide beneficial effects via anti-oxidative activity against LDL oxidation and excess oxidative stress in the blood.

Streptococcus thermophilus fermented milk reduces serum MDA-LDL and blood pressure in healthy and mildly hypercholesterolaemic adults.

Low-density lipoprotein (LDL)-cholesterol, malondialdehyde-modified low-density lipoprotein (MDA-LDL), MDA-LDL/LDL-cholesterol in serum, and blood pressure are considered useful risk markers of cardiovascular diseases. This study aimed to examine whether a fermented milk containing Streptococcus thermophilus YIT 2001 (ST), which has high anti-oxidative activity, would benefit healthy and mildly hyper-LDL-cholesterolaemic adults via a randomised, double-blind, placebo-controlled trial. ST-fermented milk or non-fermented placebo milk (PC) was consumed once a day for 12 weeks by 29 and 30 subjects, respectively, with average serum LDL-cholesterol levels of about 140 mg/dl. Serum levels of LDL-cholesterol and MDA-LDL and blood pressure were analysed before (baseline) and after consumption. Comparisons of the responses between both groups were assessed using analysis of covariance (ANCOVA, with the baseline value as the covariate). ANCOVA demonstrated that the ST group had significant reductions in MDA-LDL, MDA-LDL/LDL-cholesterol, systolic blood pressure (SBP), and diastolic blood pressure (DBP) compared with the PC group during the consumption period (P<0.05). Moreover, stratified analysis revealed that there were significant reductions in MDA-LDL, MDA-LDL/LDL-cholesterol, SBP, and DBP in the ST group compared with the PC group during the consumption period in subjects who had above median (65 U/l) levels of oxidative stress marker MDA-LDL at baseline (P<0.05), but not in subjects with levels below the median. These findings suggest that daily consumption of ST-fermented milk may be beneficial in healthy or mildly hyper-LDL cholesterolaemic subjects through reductions in risk marker values of oxidative stress and/or cardiovascular diseases. The benefits were particularly remarkable in subjects who had higher levels of MDA-LDL.

Outcomes of 23- and 25-gauge transconjunctival sutureless vitrectomies for idiopathic macular holes.

BACKGROUND/AIMS: To assess the outcomes of 23-gauge sutureless transconjunctival vitrectomies (TSV), as compared with 25-gauge TSV in macular hole surgeries. METHODS: A retrospective, consecutive, interventional case series of 47 eyes with idiopathic macular holes treated by 23- or 25-gauge TSV were analysed. RESULTS: The operative time was 37.2 (SD 8.9) min with 23-gauge TSV and 34.2 (8.7) min with 25-gauge TSV (p = 0.388). The anatomical success rate was 96% with 23-gauge TSV and 92% with 25-gauge TSV (p>0.999). The logarithm of the minimum angle of resolution of best-corrected visual acuity (BCVA) at the sixth postoperative month was 0.19 (0.16) with 23-gauge TSV and 0.19 (0.25) with 25-gauge TSV (p = 0.521). Postoperative improvement in BCVA was comparable between the two TSVs. IOP on postoperative day 1 was lower with 25-gauge TSV (12.3 (4.9) mm Hg) than with 23-gauge TSV (17.4 (5.8) mm Hg) (p = 0.036). Complications included retinal break, intraoperative bleeding and slippage of the infusion cannula with 23-gauge TSV, while retinal detachment and postoperative hypotony occurred in the 25-gauge TSV group (p = 0.570). CONCLUSION: 23-gauge TSV appears to be as safe and effective as 25-gauge TSV in macular hole surgery.

Evaluation of optic nerve head configuration in various types of optic neuropathy with Heidelberg Retina Tomograph.

PURPOSE: To test whether Heidelberg Retina Tomograph (HRT) is applicable to assess the optic nerve head (ONH) configuration of the atrophic phase of non-glaucomatous optic neuropathy when a default set of the reference plane is used. METHODS: Ten eyes with non-arteritic anterior ischaemic optic neuropathy (NAION), 17 eyes with Leber's hereditary optic neuropathy (LHON), 40 eyes with compressive optic neuropathy (CON) owing to chiasmal tumour, and 241 eyes of control individuals were examined with HRT using the default reference plane. The global values of HRT parameters were evaluated among the groups of patients and controls. The sectoral measurements of the eyes with LHON and CON were compared with controls. To eliminate the influence of disc size and age on HRT measurements, eyes with disc area- and age-matched normal controls were used for comparison with eyes with NAION and LHON. RESULTS: Cup parameters in eyes with NAION were similar to those in controls. The retinal nerve fibre layer (RNFL) thickness was significantly thinner in eyes with NAION than that of controls. The eyes with LHON had significantly larger cup parameters, smaller rim volume, and thinner mean RNFL thickness than controls. Eyes with CON had significantly larger rim area and smaller cup parameters but similar RNFL thickness compared with controls. CONCLUSIONS: When the default reference plane is used, HRT can measure the ONH configuration in eyes with NAION and LHON as expected. However, caution must be made to interpret the parameters obtained from the eyes with CON.

Expression and localisation of calbindin D28k in all intestinal segments of the laying hen.

1. The aim of the present study was to investigate expression and localisation of a 28-kDa calcium-binding protein (CaBP-D28k) related to active calcium (Ca) absorption, in the entire intestine of egg-laying hens. 2. Western blotting analysis showed that the entire intestine expressed CaBP-D28k to the following degree: duodenum > jejunum > caecum > ileum > colon. Immunohistochemistry showed strong CaBP-D28k localisation in enterocytes along the villus tip-crypt axis in the duodenum and in villus tips in the caecum and colon. The jejunum and ileum had moderate localisation with respect to the number of immunoreactive cells and staining intensity. 3. These results suggest that laying hens actively absorb Ca in both the large and small intestines.

Macular thickness reduction in eyes with unilateral optic atrophy detected with optical coherence tomography.

AIMS: To assess the changes in macular and peripapillary retinal nerve fibre layer (RNFL) thickness in eyes with unilateral optic atrophy and to evaluate the relationship between retinal thickness and visual function. METHODS: Enrolled were 22 patients with unilateral optic atrophy. Macular thickness at the divided nine areas and peripapillary RNFL thickness in quadrantic sections were measured by optical coherence tomography. Thickness values in the affected eyes were compared with those in the contralateral unaffected eyes. The correlation of foveal thickness with best-corrected visual acuity (BCVA) was evaluated. The correlation between retinal thickness and the remaining visual field area circumscribed with I-4-e isopter in superior and inferior hemifield was assessed. RESULTS: Macular thinning was observed in all areas (P < 0.001 in each area) other than the fovea (P = 0.068). Peripapillary RNFL thickness decreased in all quadrantic sections (P < 0.001 in each section). The affected to unaffected eye ratio of retinal thickness was more than 0.6 in each area. BCVA did not correlate with foveal thickness (correlation coefficient = 0.094, P = 0.668). Although not statistically significant (P = 0.281, superior hemifield; P = 0.053, inferior hemifield), there was a tendency that eyes with severe visual field loss show more marked retinal thinning. CONCLUSIONS: Macular thinning with the preserved foveal thickness is a hallmark of eyes with optic atrophy. Together with no correlation between foveal thickness and BCVA, this finding would help in differential diagnosis of macular and optic nerve diseases.

Adhesive and bone resorptive activities of isolated osteoclasts from hen medullary bone.

In the present study, osteoclasts were isolated from hen medullary bones at the formative and resorptive phases. The cells were cultured on glass culture dishes and bone slices. After culturing, the adhesion activity of the isolated osteoclasts with the substrates was estimated with a light microscope, and the surfaces of the bone slices were observed with a scanning electron microscope. The results showed that the adhesion activity of tartrate-resistant acid phosphatase (TRAP)-positive osteoclasts is higher at the bone resorptive phase than at the bone formative phase, and this tendency in isolated osteoclasts was observed more frequently on the bone slices than on the glass culture dishes. Furthermore, scanning electron microscopy showed that the isolated osteoclasts in the bone resorptive phase adhered to the bone surface with developed-cytoplasmic projections and formed broad pits where collagen fibrils were exposed. On the other hand, isolated osteoclasts in the bone formative phase adhered to the bone slice with board-shaped cytoplasmic projections and did not form any pits. These results suggest that isolated osteoclasts in the bone resorptive phase have a high level of adhesion activity and actively resorb the bone, whereas isolated osteoclasts in the bone formative phase have a low level of adhesion activity and cease bone resorption. The procedure reported here is useful for studying the bone-resorptive mechanism of authentic osteoclasts.

The kinetics of actin filaments in osteoclasts on chicken medullary bone during the egg-laying cycle.

Distributions of actin filaments in osteoclasts on chicken medullary bone were examined during the egg-laying cycle. When an egg was in the magnum or isthmus of the oviduct at 6 h after oviposition, the osteoclasts lacked ruffled borders and attached to medullary bone surface via clear zones. Actin filaments were localized as amorphous bands in the bone site of the cytoplasm, corresponding to clear zones in the osteoclasts. When an egg was in the shell gland of the oviduct at 15 h after oviposition, the osteoclasts had long and slender, well-developed ruffled borders. Actin filaments were localized as lines perpendicular to the bone surface at the bone site of the cytoplasm. In medullary bone osteoclasts, actin filaments changed during the egg-laying cycle. The relationship between the actin filaments and the ultrastructure of the osteoclasts are discussed in this report.

Ultrastructural changes of osteoclasts on hen medullary bone during the egg-laying cycle.

1. The ultrastructure of osteoclasts on hen medullary bone during the egg-laying cycle was observed by electron microscopy. 2. At 0 to 6 h after oviposition, osteoclasts lacking ruffled borders were attached to the bone via the clear zone and appeared to have ceased bone resorption. Small vacuoles were scattered throughout this cytoplasm. 3. At 9 to 21 h after oviposition, most of the osteoclasts had ruffled borders and appeared to be resorbing bone. The ruffled borders at 15 h were well developed, whereas at 9, 18 and 21 h they were poorly developed or showed similarities in structure to the clear zone. The small vacuoles were concentrated under the ruffled borders at 9 h and then decreased. 4. These results demonstrate cyclic changes in osteoclasts during the egg-laying cycle, indicate that ruffled borders form at the beginning of bone resorption and suggest that they are derived from the clear zone and the small vacuoles. The results also indicate that the ruffled borders fuse into the clear zone and disappear at the completion of bone resorption.

Immunoelectron microscopic detection of estrogen target cells in the bone marrow of estrogen-treated male Japanese quail.

The localization of estrogen receptors (ERs) in bone marrow cells was studied immunoelectron microscopically in the femurs of estrogen-treated male Japanese quail. The electron-dense reaction product showing ER localization was not detected in the bone marrow cells of the male quail. After 48 h of estrogen treatment, nuclear reaction product was observed in many mononuclear cells in the bone marrow. Most were located in the marrow stroma, and they were divided into undifferentiated and differentiated type cells, characterized by poorly and well-developed cytoplasm, respectively. Some ER-positive cells were situated in the hematopoietic tissue and had moderately developed cytoplasm. Thus, estrogen target cells may be present in the stroma and hematopoietic tissue of bone marrow.

Estrogen target cells during the early stage of medullary bone osteogenesis: immunohistochemical detection of estrogen receptors in osteogenic cells of estrogen-treated male Japanese quail.

The localization of estrogen receptors (ER) in osteogenic cells during the early stage of medullary bone osteogenesis was studied immunohistochemically in the femurs of estrogen-treated male Japanese quail. Alkaline phosphatase (ALP) activity was used as a marker for osteogenic cells. ER immunostaining was observed in the nuclei of weak ALP-positive bone lining cells on the endosteal bone surface of nontreated birds. After 24 hours of estrogen treatment, nuclear immunostaining was detected in ALP-positive preosteoblasts on the endosteal bone surface. After 48 hours, the medullary bone appeared to some degree along the endosteal surface. ER immunostaining was observed in the nuclei of ALP-positive osteoblasts on the medullary bone surface. This study demonstrates that ER are present in osteogenic cells, and suggests that estrogen directly acts on medullary bone osteogenesis.

Immunoelectron microscopic demonstration of estrogen receptors in osteogenic cells of Japanese quail.

The localization of estrogen receptors (ERs) in osteogenic cells was immunoelectron microscopically examined in the femurs of female and estrogen-treated male Japanese quail. An electron dense reaction product showing ER localization was observed in the nuclei of osteoblasts and immature osteocytes in the medullary bone of the female quail. However, reaction product was not seen in the osteoclasts. On the endosteal bone surface of male quail, nuclear reaction product was detected in bone lining cells. After 24 h of estrogen treatment, reaction product was observed in the nuclei of preosteoblasts on the endosteal bone surface. After 48 h, the medullary bone partly appeared along the endosteal surface. Nuclear reaction product was seen in osteoblasts on the medullary bone surface.

Histochemical identification of oestrogen target cells in the medullary bone of laying hens.

1. Oestrogen target cells in the medullary bone of laying hens were examined using a histochemical method with fluorescein isothiocyanate labelled oestradiol. 2. Specific fluorescence was found to occur strongly in osteoblasts and weakly in osteoclasts on the medullary bone surface. 3. This study suggests that osteoblasts on the medullary bone surface are oestrogen target cells.

Effects of oestrogen and anti-oestrogen on the cells of the endosteal surface of male Japanese quail.

1. The endosteal cellular response occurring in the early stages of medullary bone formation was histologically examined in the femurs of male Japanese quail treated with oestrogen and antioestrogen. 2. The numbers of endosteal cells decreased, and preosteoblasts and osteoblasts increased, on the endosteal surface following oestrogen treatment. 3. After oestrogen and anti-oestrogen treatment the numbers of preosteoblasts and osteoblasts on the endosteal surface did not increase. However, endosteal cells slowly decreased in number and osteoclasts increased. 4. This study demonstrates that differentiation of osteoblasts is induced by oestrogen stimulation and suggests that osteoclasts occur by oestrogen inhibition on the endosteal surface.

Cytology and autoradiography of estrogen-induced differentiation of avian endosteal cells.

The endosteal reaction, the initial step in the formation of medullary bone, was investigated in femurs of estrogen-treated male Japanese quail. Morphologically, the endosteal cells were in an undifferentiated state until 30 h after estrogen treatment and showed characteristics resembling those of resting cells. Many preosteoblasts were seen on the endosteum at 33 h, whereas mitotic figures and fully differentiated osteoblasts were recognized at 36 h after estrogen. The mitotic figures were observed among the preosteoblasts on the endosteum. Autoradiographs showed that the number of endosteal cells labeled by [3H]thymidine injected 1 h before sacrifice was maximal 27 h after the estrogen administration and decreased markedly by 30 h. When a single injection of [3H]thymidine was given at 26 h after estrogen, the highest percent of labeled endosteal cells was observed 1 h later (27 h after estrogen). Labeled preosteoblasts and osteoblasts were observed at 7 h (33 h after estrogen) and 10 h (36 h after estrogen), respectively. Our results show that under the influence of estrogen, endosteal cells are induced to maximally synthesize DNA about 27 h after estrogen. These cells appear to modulate into preosteoblasts in about 6 h and then divide via mitosis to become osteoblasts within an additional 3 h. The development of medullary bone induced by estrogen occurs in a sequential and predictable manner, which makes it a useful system for studying basic problems on bone cell differentiation.

Xanthic variant of non-ossifying fibroma (so-called xanthofibroma) of the mandible. An ultrastructural study.

An unusual case of a 37-year-old female with xanthomatous bone tumor of the right molar area of the mandible was presented. The tumor was asymptomatic and found to be a well-demarcated intraosseous radiolucent lesion on radiographic examination. Histologically the tumor consisted of two cell types, fibroblastic and xanthomatous cells. There was no osteoid, bone or cartilage formation. However, numerous psammomatous calcified bodies were seen in the fibrous area. Ultrastructural study showed fibroblastic cells in different stages of proliferation as the basis of the tumor which transform itself into xanthomatous cells. From the clinicopathologic findings, our case was thought to be a xanthic variant of non-ossifying fibroma (so-called xanthofibroma) of the mandible.