The effect of femoral bone tunnel configuration on tendon-bone healing in an anterior cruciate ligament reconstruction: An animal study.

OBJECTIVES: To compare the effect of femoral bone tunnel configuration on tendon-bone healing in an anterior cruciate ligament (ACL) reconstruction animal model. METHODS: Anterior cruciate ligament reconstruction using the plantaris tendon as graft material was performed on both knees of 24 rabbits (48 knees) to mimic ACL reconstruction by two different suspensory fixation devices for graft fixation. For the adjustable fixation device model (Socket group; group S), a 5 mm deep socket was created in the lateral femoral condyle (LFC) of the right knee. For the fixed-loop model (Tunnel group; group T), a femoral tunnel penetrating the LFC was created in the left knee. Animals were sacrificed at four and eight weeks after surgery for histological evaluation and biomechanical testing. RESULTS: Histologically, both groups showed a mixture of direct and indirect healing patterns at four weeks, whereas only indirect healing patterns were observed in both groups at eight weeks. No significant histological differences were seen between the two groups at four and eight weeks in the roof zone (four weeks, S: mean 4.8 sd 1.7, T: mean 4.5 sd 0.5, p = 0.14; eight weeks, S: mean 5.8 sd 0.8, T: mean 4.8 sd 1.8, p = 0.88, Mann-Whitney U test) or side zone (four weeks, S: mean 5.0 sd 1.2, T: mean 4.8 sd 0.4, p = 0.43; eight weeks, S: mean 5.3 sd 0.8,T: mean 5.5 sd 0.8, p = 0.61, Mann-Whitney U test) . Similarly, no significant difference was seen in the maximum failure load between group S and group T at four (15.6 sd 9.0N and 13.1 sd 5.6N) or eight weeks (12.6 sd 3.6N and 17.1 sd 6.4N, respectively). CONCLUSION: Regardless of bone tunnel configuration, tendon-bone healing after ACL reconstruction primarily occurred through indirect healing. No significant histological or mechanical differences were observed between adjustable and fixed-loop femoral cortical suspension methods.Cite this article: Y. Sato, R. Akagi, Y. Akatsu, Y. Matsuura, S. Takahashi, S. Yamaguchi, T. Enomoto, R. Nakagawa, H. Hoshi, T. Sasaki, S. Kimura, Y. Ogawa, A. Sadamasu, S. Ohtori, T. Sasho. The effect of femoral bone tunnel configuration on tendon-bone healing in an anterior cruciate ligament reconstruction: An animal study. Bone Joint Res 2018;7:327-335. DOI: 10.1302/2046-3758.75.BJR-2017-0238.R2.

The effect of systemic administration of G-CSF on a full-thickness cartilage defect in a rabbit model MSC proliferation as presumed mechanism: G-CSF for cartilage repair.

OBJECTIVES: The aim of this study was to investigate the effect of granulocyte-colony stimulating factor (G-CSF) on mesenchymal stem cell (MSC) proliferation in vitro and to determine whether pre-microfracture systemic administration of G-CSF (a bone marrow stimulant) could improve the quality of repaired tissue of a full-thickness cartilage defect in a rabbit model. METHODS: MSCs from rabbits were cultured in a control medium and medium with G-CSF (low-dose: 4 µg, high-dose: 40 µg). At one, three, and five days after culturing, cells were counted. Differential potential of cultured cells were examined by stimulating them with a osteogenic, adipogenic and chondrogenic medium.A total of 30 rabbits were divided into three groups. The low-dose group (n = 10) received 10 µg/kg of G-CSF daily, the high-dose group (n = 10) received 50 µg/kg daily by subcutaneous injection for three days prior to creating cartilage defects. The control group (n = 10) was administered saline for three days. At 48 hours after the first injection, a 5.2 mm diameter cylindrical osteochondral defect was created in the femoral trochlea. At four and 12 weeks post-operatively, repaired tissue was evaluated macroscopically and microscopically. RESULTS: The cell count in the low-dose G-CSF medium was significantly higher than that in the control medium. The differentiation potential of MSCs was preserved after culturing them with G-CSF.Macroscopically, defects were filled and surfaces were smoother in the G-CSF groups than in the control group at four weeks. At 12 weeks, the quality of repaired cartilage improved further, and defects were almost completely filled in all groups. Microscopically, at four weeks, defects were partially filled with hyaline-like cartilage in the G-CSF groups. At 12 weeks, defects were repaired with hyaline-like cartilage in all groups. CONCLUSIONS: G-CSF promoted proliferation of MSCs in vitro. The systemic administration of G-CSF promoted the repair of damaged cartilage possibly through increasing the number of MSCs in a rabbit model.Cite this article: T. Sasaki, R. Akagi, Y. Akatsu, T. Fukawa, H. Hoshi, Y. Yamamoto, T. Enomoto, Y. Sato, R. Nakagawa, K. Takahashi, S. Yamaguchi, T. Sasho. The effect of systemic administration of G-CSF on a full-thickness cartilage defect in a rabbit model MSC proliferation as presumed mechanism: G-CSF for cartilage repair. Bone Joint Res 2017;6:123-131. DOI: 10.1302/2046-3758.63.BJR-2016-0083.

The relationship between oxygen consumption rate and viability of in vivo-derived pig embryos vitrified by the micro volume air cooling method.

The aim of this study was to assess the viability of vitrified-warmed in vivo-derived pig embryos after measuring the oxygen consumption rate. Six days after artificial insemination, blastocysts were collected from gilts and vitrified by the micro volume air cooling method. The oxygen consumption rate was measured in 60 vitrified-warmed embryos, which were then cultured for 48h to assess the viability. The survival (re-expansion) rate of embryos after warming was 85.0%. The average oxygen consumption rate of embryos immediately after warming was greater in embryos which could re-expand during subsequent culture (F=0.75±0.04) than that in those which failed to re-expand (F=0.33±0.05). Moreover, the oxygen consumption rate of vitrified-warmed embryos was greater in the hatched (F=0.88±0.06) than that in the not-hatched group (F=0.53±0.04). When the oxygen consumption rate of the vitrified-warmed embryos and the numbers of viable and dead cells in embryos were determined, there was a positive correlation between the oxygen consumption rate and the number of live cells (P<0.01, r=0.538). A total of 29 vitrified embryos after warming and measuring the oxygen consumption rate were surgically transferred into uterine horns of two recipients. Both of the recipients become pregnant and farrowed 12 healthy piglets. These results demonstrate that the oxygen consumption rate of vitrified-warmed pig embryos can be related to the number of live cells and that the measurement of oxygen consumption of embryos after cryopreservation may be useful for estimating embryo survivability.

Relationships between oxygen consumption rate, viability, and subsequent development of in vivo-derived porcine embryos.

Oxygen consumption rate of in vivo-derived porcine embryos was measured, and its value as an objective method for the assessment of embryo quality was evaluated. Embryos were surgically collected 5 or 6 days after artificial insemination (AI), and oxygen consumption rate of embryos was measured using an embryo respirometer. The average oxygen consumption rate (F × 10(14)/mol s(-1)) of the embryos that developed to the compacted morula stage on Day 5 (Day 0 = the day of artificial insemination) was 0.58 ± 0.03 (mean ± standard error of the mean). The Day-6 embryos had consumption rates of 0.56 ± 0.13, 0.87 ± 0.06, and 1.13 ± 0.07 at the early blastocyst, blastocyst, and expanded blastocyst stages, respectively, showing a gradual increase as the embryos developed. Just after collection, the average oxygen consumption rates of embryos that hatched and of those that did not hatch after culture were 0.60 ± 0.04 and 0.50 ± 0.04 for Day 5 (P = 0.08) and 1.05 ± 0.09 and 0.77 ± 0.05 for Day 6 (P < 0.05), respectively. The value and probability of discrimination by measuring the oxygen consumption rates of embryos to predict their hatching ability after culture were 0.56 and 63.6% for Day-5 embryos and 0.91 and 68.4% for Day-6 blastocysts, respectively. When Day-5 embryos were classified based on the oxygen consumption rate and then transferred non-surgically to recipient sows, three of the seven sows, to which embryos having a high oxygen consumption rate (≥ 0.59) were transferred, became pregnant and farrowed a total of 20 piglets. However, none of the four sows, to which embryos having low oxygen consumption rate (< 0.59) were transferred, became pregnant. These results suggest that the viability of in vivo-derived porcine embryos and subsequent development can be estimated by measuring the oxygen consumption rate.

Hidden osteophyte formation on plain X-ray is the predictive factor for development of knee osteoarthritis after 48 months--data from the Osteoarthritis Initiative.

OBJECTIVE: To examine whether the detection of osteophytes anywhere in the knee could serve as a pre-radiographic biomarker for osteoarthritis (OA) development. METHODS: Baseline magnetic resonance imaging (MRIs) of 132 participants in the Osteoarthritis Initiative (OAI) were studied. Based on radiographs, 66 knees were assessed as osteoarthritis-free (no-osteoarthritis [NOA], or Kellgren/Lawrence [K/L] severity grade 0/1 both at baseline and 48 months), and another 66 knees were assessed as having radiographic OA changes (pre-radiographic osteoarthritis [PROA], or with K/L grade 0/1 at baseline and grade ≥ 2 at 48 months). Using baseline MRI data, we examined eight sites of osteophyte formation: the medial and lateral femoral condyle (MFC and LFC, respectively); medial and lateral tibial plateau (MTP and LTP, respectively); medial and lateral facets of the patellofemoral joint (PM and PL, respectively); tibial spine (TS); and femoral intercondylar notch (IC). Knee joint osteophyte size was assessed via the 8-point marginal osteophytes item of the whole-organ magnetic resonance imaging score (WORMS). The frequencies and distributions of osteophytes were compared between groups. RESULTS: Mild-size osteophytes (defined as score ≥ 2) were observed more frequently at the MFC (P = 0.00278), MTP (P = 0.0046), TS (P = 0.0146), PM (P < 0.0001), PL (P = 0.0012), and IC (P < 0.0001) in PROA knees than in NOA knees. Moderate-size osteophytes (defined as score ≥ 4) were more frequently observed in PROA knees than in NOA knees only at the IC (P < 0.0001). CONCLUSION: Knees with osteophyte formation at the IC, even those of K/L severity grade 0/1, are at risk for the development of radiographic OA by 48 months.

Preventive effects of hyaluronan from deterioration of gait parameters in surgically induced mice osteoarthritic knee model.

OBJECTIVE: Osteoarthritis (OA) leads to pain and loss of function in affected joints. Gait disturbance results from these symptoms of OA, and gait analysis can be important to evaluate the progression of OA. The purpose of this study was to analyze gait pattern in a rodent model of OA and to assess the effects of intra-articular injection of hyaluronan (IAI-HA) by gait analysis, along with histological evaluation. DESIGN: OA was induced by destabilization of the medial meniscus (DMM) of C57BL/6 mice. IAI-HA started 3 weeks after DMM surgery. Mice were allocated to three groups and were given either 800-kDa HA (800-HA), 6000-kDa HA (6000-HA) or saline. We compared these three groups with a sham group by gait analysis using CatWalk. Histological evaluation was performed to assess articular cartilage changes in the knee joints. RESULTS: Mice injected with 800-HA or 6000-HA showed gait patterns similar to that of the sham mice, while the saline-injected group showed gait disturbances 12 and 16 weeks after DMM surgery. Histological changes in articular cartilage were similar among the 800-HA, 6000-HA and saline-treated groups, demonstrating OA progression throughout the experimental time points. Positive gait-related effects of IAI-HA might occur by its pain relieving effect and/or by preventing contracture. CONCLUSION: IAI-HA prevented gait disturbances in the DMM model, but did not prevent histological changes associated with OA progression.

Cerebral cyst formation following stereotactic ablative irradiation for non-nasopharyngeal head and neck malignancies: imaging findings and relevant dosimetric parameters.

OBJECTIVE: To describe the clinical characteristics, imaging findings and relevant dosimetric parameters of cases presenting with cerebral cyst formation (CCF) after single or oligo-fractionated stereotactic radiotherapy (SRT) for non-nasopharyngeal head and neck malignancies (HNMs). METHODS: We identified four cases with the follow-up duration of 5.7-9.1 years from SRT. The irradiated sites included the middle ear in one case and the ethmoid sinus in three cases, two of the latter possessed brain invasion. The chronological changes in MR images and the dose-volume histogram of the adjacent brain tissue were evaluated. RESULTS: CCF with or without multiple septi presented with a latency of 29-86 months (median, 45.5 months), which was preceded by either non-specific parenchymal enhancement or typical radiation necrosis. In three cases, CCF adjacent to the frontal base resultantly caused mass effect, and two of these three cases required surgical intervention at 38 and 54 months, respectively, after SRT for alleviation of symptoms. The relation of the irradiated brain volumes to the biological equivalent dose based on the linear-quadratic (LQ) and LQ-cubic models was represented as a threshold. CONCLUSION: When contemplating SRT for HNM cases, caution should be exercised to the dose-volume relation-ship of the adjacent brain tissue, especially the frontal base, as well as other critical structures, and long-term vigilant follow-up is also mandatory. ADVANCES IN KNOWLEDGE: CCF can occur as an unusual consequence of late brain injury with variable but mostly long latency following SRT for non-nasopharyngeal HNMs adjacent to the brain, even superficial parts that were previously irradiated via conventional radiotherapy.

Consideration of optimal isodose surface selection for target coverage in micro-multileaf collimator-based stereotactic radiotherapy for large cystic brain metastases: comparison of 90%, 80% and 70% isodose surface-based planning.

OBJECTIVE: This study aims to compare dynamic conformal arc (DCA) plans based on different-percentage isodose surfaces (IDSs), normalised to 100% at the isocentre, for target coverage (TC; dose prescription) in stereotactic radiotherapy for large cystic brain metastases. METHODS: The DCA plans were generated for 15 targets (5 spherical models and 10 metastatic brain lesions) based on 90%, 80% and 70% IDSs for dose prescription to attain ≥99% TC values using the Novalis Tx platform. These plans were optimised mainly by leaf margin and/or collimator angle adjustment, while similar arc arrangements were used. RESULTS: TC values were equivalent among the three plans. Conformity index values were similar between the 80% and 70% plans, while they were worse in the 90% plans. Mean doses (D(mean)) of the interior 3 mm rind structure were highest in the 70% plans, followed by the 80% plans and lowest in the 90% plans. D(mean) of the exterior 3 mm rind structure and the ratio of 50%/100% isodose volumes (Paddick's gradient index values) were highest in the 90% plans, followed by 80% and lowest in the 70% plans. CONCLUSIONS: These results suggest that the 70% IDS plans might be beneficial for both tumour control and reducing toxicity to surrounding normal tissue if appropriate dose conformity and precise treatment set-up are ensured. The 90% IDS plans are unfavourable in view of inferior dose gradient outside the target and should be limited to cases in which the target dose homogeneity is given the highest priority.

The relation between various conformity indices and the influence of the target coverage difference in prescription isodose surface on these values in intracranial stereotactic radiosurgery.

OBJECTIVE: The purpose of this study was to describe the relation between various frequently used conformity indices (CIs) and to examine the influence of the target coverage (TC) difference in prescription isodose surface (IDS) on these CI values in dynamic conformal arc (DCA) plans. METHOD: 73 plans for simple-shaped brain metastases that were previously characterised for dose distribution with regard to the effect of the target volume (TV) and the depth from the skin surface were reviewed. Three different-definition CI values for each TV were calculated at the 80% IDS, and at D99, D95, D90 and D85, considering the interplanner variability in the TC values for the prescription IDS. RESULTS: The CI used as the Radiation Therapy Oncology Group criterion showed nearly perfect values at D90. The CI defined in the BrainSCAN (BrainLAB AG, Feldkirchen, Germany) treatment planning system (CI(BS)) denoted lower (superior) values as the TC of the reference IDS decreased. Nakamura's CI (NCI) had lower variability but demonstrated lower (superior) values at D95. NCI showed the most stringent (higher) values at an 80% IDS, but the differences between the plans were less distinct with NCI. CONCLUSION: The TC difference in IDS chosen for dose prescription or evaluation significantly led to CI value variability in a definition-dependent manner, even when NCI was applied. Definition of the reference IDS at a specific TC value according to clinical situation would reduce the CI value variability to a minimum and would make the CI(BS) sufficient for the objective metric with a perfect value of 1.

Characterisation of dose distribution in linear accelerator-based intracranial stereotactic radiosurgery with the dynamic conformal arc technique: consideration of the optimal method for dose prescription and evaluation.

OBJECTIVES: The purpose of this study was to characterise dose distribution in linear accelerator-based intracranial stereotactic radiosurgery using the dynamic conformal arc technique, and to validate the pertinence of dose prescription to the specific percentage isodose surface (IDS). METHODS: 73 plans for brain metastases were reviewed and replanned with a uniform method for target definition and treatment planning. RESULTS: In all cases except 1 the dose prescription to the 80% IDS satisfied the criteria of the standardised prescription IDS as previously proposed. However, both of the planning target volume (PTV) coverage values for the 80% and 90% IDSs and the PTV D99 and D95 (IDS receiving at least 99% or 95% of the PTV) were inconsistent and significantly increased as a function of the PTV size. The 80% IDS for a PTV of more than 5 cm(3) achieved adequate PTV coverage without a leaf margin. The dose conformity for 80% IDS gradually worsened as the PTV increased, whereas that for the PTV D99 or D95 improved as a function of the PTV size. The addition of a leaf margin attained 100% PTV coverage for 80% IDS, while leading to a poorer dose conformity. CONCLUSION: The dose prescription to the specific percentage IDS does not necessarily guarantee consistent target coverage, D99 and D95, and desirable dose conformity in proportion to the target volume. The dose prescription and evaluation at the specific target coverage would therefore be preferable as an objective method in order to report the "marginal dose" and to clearly compare the planning parameters with those from other modalities.

Pro-apoptotic activity of inhibitory PAS domain protein (IPAS), a negative regulator of HIF-1, through binding to pro-survival Bcl-2 family proteins.

Inhibitory PAS (Per/Arnt/Sim) domain protein (IPAS) is a dominant negative transcription factor that represses hypoxia-inducible factor 1 (HIF-1) activity. In this study, we show that IPAS also functions as a pro-apoptotic protein through binding to pro-survival Bcl-2 family members. In a previous paper, we reported that NF-κB-dependent IPAS induction by cobalt chloride repressed the hypoxic response in PC12 cells. We found that prolonged incubation under the same conditions caused apoptosis in PC12 cells. Repression of IPAS induction protected cells from apoptosis. Furthermore, knockdown of IPAS recovered cell viability. EGFP-IPAS protein was localized in both the nucleus and the cytoplasm, with a large fraction associated with mitochondria. Mitochondrial IPAS induced mitochondria depolarization and caspase-3 activation. Immunoprecipitation assays revealed that IPAS is associated with Bcl-x(L), Bcl-w and Mcl-1. The association of IPAS with Bcl-x(L) was also observed in living cells by the FLIM-based FRET analysis, indicating direct binding between the two proteins. IPAS contributed to dysfunction of Bcl-x(L) by inhibiting the interaction of Bcl-x(L) with Bax. These results demonstrate that IPAS functions as a dual function protein involved in transcription repression and apoptosis.

SS-MIX: a ministry project to promote standardized healthcare information exchange.

OBJECTIVES: To promote healthcare information exchange between providers and to allow hospital information systems (HIS) export information in standardized format (HL7 and DICOM) in an environment of wide-spread legacy systems, which only can export data in proprietary format. METHODS: Through the Shizuoka prefecture EMR project in 2004-2005, followed by the ministry's SS-MIX project, many software products have been provided, which consist of 1) a standardized storage to receive HL7 v2.5 messages of patient demographics, prescription orders, laboratory results, and diagnostic disease in ICD-10, 2) a referral letter creation system, 3) a formatted document creation system, 4) a progress note/nursing record system, and 5) an archive/viewer to incorporate incoming healthcare data CD and allow users to view on HIS terminal. Meanwhile, other useful applications have been produced, such as adverse event reporting and clinical information retrieval. To achieve the above-mentioned objectives, these software products were created and propagated, because users can use these software products, provided that their HIS can export the above information to the standardized storage in HL7 v2.5 format. RESULTS: In 20 hospitals of Japan, the standardized storage has been installed and some applications have been used. As major HIS vendors are shipping HIS with HL7 export function since 2007, HIS of 594 hospitals in Japan became capable of exporting data in HL7 v2.5 format (as of March 2010). CONCLUSIONS: In high CPOE installation rate (85% in 400+ bed hospitals), though most of them only capable of exporting data in proprietary format, prefecture and ministry projects were effective to promote healthcare information exchange between providers. The standardized storage became an infrastructure for many useful applications, and many hospitals started using them. Ministry designation of proposed healthcare standards was effective so as to allow vendors to conform their products, and users to install them.

Automated segmentation of recuts abdominis muscle using shape model in X-ray CT images.

Our purpose in this study is to segment the rectus abdominis muscle region in X-ray CT images, and we propose a novel recognition method based on the shape model. In this method, three steps are included in the segmentation process. The first is to generate a shape model for the rectus abdominis muscle. The second is to recognize anatomical feature points corresponding to the origin and insertion of the muscle, and the third is to segment the rectus abdominis muscles based on the shape model. We generated the shape model from 20 CT cases and tested the model to recognize the muscle in 20 other CT cases. The average values for the Jaccard similarity coefficient (JSC) and true segmentation coefficient (TSC) were 0.841 and 0.863, respectively. The results suggest the validity of the model-based segmentation for the rectus abdominis muscle.

Automated recognition of the psoas major muscles on X-ray CT images.

The purpose of this study is to recognize the psoas major muscle on X-ray CT images. For this purpose, we propose a novel recognition method. The recognition process in this method involves three steps: the generation of a shape model for the psoas major muscle, recognition of anatomical points such as the origin and insertion, and the recognition of the psoas major muscles by the use of the shape model. We generated the shape model using 20 CT cases and tested the model for recognition in 20 other CT cases. The average Jaccard similarity coefficient (JSC) and reproducibility rate were 0.704 and 0.783, respectively. Experimental results indicate that our method was effective for a 2-D cross-sectional area (CSA) analysis.

Automated measurement of bone-mineral-density (BMD) values of vertebral bones based on X-ray torso CT images.

Bone is one of the most important anatomical structures in humans and osteoporosis is one of the major public health concerns in the world. Osteoporosis is a main target disease of bone, which can be detected by medical image techniques. The purpose of this study is to develop a fully automated computer scheme to measure bone-mineral-density (BMD) values for vertebral trabecular bones. This scheme will aid osteoporosis diagnosis performed using computer tomography (CT) images. This scheme includes the following processing steps: segmentation of the bone region, recognition of the skeletal structures and measurement of the BMD value in vertebral trabecular bone of each vertebral body. The proposed scheme was applied to 20 X-ray torso CT cases to measure the BMD values for vertebral trabecular bones. The experimental results show that the mean and standard deviation of the difference between the BMD values measured by using the proposed method and those measured using a manual segmentation method were 6.93 mg/cm(3) and 6.82 mg/cm(3) respectively. The accuracy of the proposed scheme satisfied the requirement for a computer-aided system used in osteoporosis diagnosis.

Transforming growth factor-alpha in a defined medium during in vitro maturation of porcine oocytes improves their developmental competence and intracellular ultrastructure.

We examined the effects of transforming growth factor-alpha (TGF-alpha) to develop a defined medium for in vitro maturation (IVM) of porcine (Sus scrofa domesticus) oocytes. Cumulus-oocyte complexes (COCs) were matured in porcine oocyte medium containing 3mg/mL polyvinyl alcohol (POM) and TGF-alpha (0, 1, 10, or 100 ng/mL) in the presence or absence of the gonadotropins equine chorionic gonadotropin (eCG) and human chorionic gonadotropin (hCG). In the absence of gonadotropins, adding 10 ng/mL TGF-alpha increased (P<0.05) the percentage of oocytes that reached metaphase II (24.2%) compared with that of the control (no TGF-alpha addition; 5.6%). In the presence of gonadotropins, although maturation rate did not differ among TGF-alpha treatments (75.4% to 84.8%), the rate of blastocyst formation (28.1%) was higher (P<0.05) in the TGF-alpha group (28.1%) than that in the control group (15.9%) after in vitro fertilization and embryo culture. An electron microscope study revealed that TGF-alpha-treated oocytes contained more homogenous lipid droplets than did control oocytes. Moreover, mitochondria surrounded by the endoplasmic reticulum were observed only in the TGF-alpha-treated oocytes. In blastocysts derived from the latter oocytes, mitochondria with numerous cristae were frequently observed compared with that in blastocysts from control oocytes. When the Day-5 blastocysts obtained from oocytes matured with TGF-alpha were surgically transferred into four recipients, a total of 29 piglets were farrowed. We concluded that the addition of TGF-alpha to the defined IVM medium of porcine oocytes improved the subsequent blastocyst formation and that the blastocysts produced by the defined in vitro production system have developmental competence to full term after embryo transfer.

Morphometric and ultrastructural changes in ciliated cells of the oviductal epithelium in prolific Chinese Meishan and Large White pigs during the oestrous cycle.

There are some differences in reproductive features between the Chinese Meishan (CM) pig and Large White (LW) pig. The aim of the present study was to investigate the quantitative changes and ultrastructural features of ciliated cells in the various regions of the CM and LW pig oviduct during the follicular and luteal phases of the oestrous cycle. In the fimbrial and ampullar epithelia at the follicular phase, the ciliated cells were more plentiful than in the isthmic region in both pigs. In the CM pigs, there was a striking decrease in the percentage and cell height of ciliated cells in the luteal phase compared with the follicular phase. Although similar quantitative changes were observed in the LW pig oviduct, these changes were less dramatic than that in the CM pig oviduct. In both pigs, the percentage and cell height of ciliated cells in the isthmus were unchanged between the follicular and luteal phases. The ultrastructure of ciliogenic and ciliated cells was observed. In the fimbrial and ampullar epithelia during the follicular phase, most of the ciliated cells showed normal morphology, having many elongated cilia and mitochondria, but in the CM pig oviduct during the luteal phase many of the ciliated cells possessed immature cilia and swollen mitochondria. Cells undergoing ciliogenesis were frequently observed in the fimbriae and ampulla, and occasionally in the isthmus. Cytoplasmic protrusions containing variable numbers of ciliary axonemal complexes occurred in the fimbrial and ampullar epithelium in the CM pigs at the luteal phase, suggesting that deciliation of cells occurs by membrane-bound cilia packets forming at the apices of cells and pinching off. These results demonstrate that there are regional variations in the cyclic changes associated with the oviductal ciliated cells of the pigs, while there are marked morphometrical and ultrastructural changes in oviductal ciliated cells of the CM pigs compared with that of the LW pigs.

Ossifying pleomorphic adenoma of the parotid gland: a case report and review.

Pleomorphic adenoma is the most common parotid gland neoplasm and occasionally arises from the deep lobe of the parotid gland with extension into the parapharyngeal space. However, ossification of pleomorphic adenoma is rare, although chondroid and myxochondroid tissue formation is frequently seen histologically. We report an unusual case of ossifying pleomorphic adenoma arising from the deep lobe of the parotid gland in the parapharyngeal space.

Regional and cyclic variations in the ultrastructural features of secretory cells in the oviductal epithelium of the Chinese Meishan pig.

The Chinese Meishan pig is prolific breed and it is considered that this pig has a capacity with higher rates of embryonic survival. The oviductal secretory cells may affect the embryonic development and survival. The aim of the present study was to investigate the ultrastructural features of secretory cells in the various regions of the Chinese Meishan pig oviduct during the follicular and luteal phases of the oestrous cycle. In the ampullar secretory cells, numerous secretory granules with moderately electron-dense matrices were present in the supranuclear cytoplasm and exocytosis of secretory granules was observed. The number of secretory granules was dramatically reduced in the ampullar secretory cells in the luteal phase. During the follicular phase in the fimbrial epithelium, the secretory cells contained rough endoplasmic reticulum and Golgi apparatus, but most cells had few small granules. In the luteal phase, the secretory cells in the ampullar and fimbrial epithelia extended beyond the luminal border of the ciliated cells. In the isthmus, many granules were present in the cytoplasm of secretory cells throughout the oestrous cycle, but the number of secretory granules was reduced in the luteal phase. The cytomorphometric data revealed that the height of ciliated cells decreased substantially in the fimbriae and ampulla at the luteal phase, while that of non-ciliated cells was less affected. These results suggest that the drastic reduction of cell height of ciliated cells cause the extrusion of most secretory cells beyond the ciliated cells in the fimbriae and ampulla during the luteal phase. In summary, our ultrastructural observations of Chinese Meishan pig oviduct revealed marked cyclic changes in the ultrastructural features of secretory cells. In particular, the ultrastructural features and the numbers of secretory granules were distinctive for each particular segment. These findings should provide insight into the regional and cellular differences in functions of secretory cells of the Chinese Meishan pig oviduct.

Characterization of green fluorescent protein-expressing retinal cells in CD 44-transgenic mice.

Sensory information in the retina is transferred from rod and cone photoreceptors to higher visual centers via numerous parallel circuits that sample the photoreceptor mosaic independently. Each circuit consists of a unique combination of ganglion cell, bipolar and amacrine cell types. The morphology and physiological responses of many amacrine cells have been characterized. However, the synaptic connections and retinal circuits in which they participate are only rarely understood. A major problem that has prevented fuller characterization of retinal circuitry is the need for specific cellular markers for the more than 50 inner retinal cell types. One potential strategy for labeling cells is to use transgenic expression of a reporter gene in a specific cell type. In a recent study of cluster of differentiation 44 (CD44)-enhanced green fluorescent protein (EGFP) transgenic mice, we observed that the green fluorescent protein (GFP) was expressed in a population of amacrine and ganglion cells in the inner nuclear layer (INL) and the GCL. To characterize the morphology of the GFP-labeled cells, whole mount preparations of the retina were used for targeted iontophoretic injections of Lucifer Yellow and Neurobiotin. Furthermore, immunocytochemistry was used to characterize the antigenic properties of the cells. We found that many GFP-expressing cells were GABAergic and also expressed calretinin. In addition to the somatic staining, there was a strong GFP(+)-band located about 50-60% depth in the inner plexiform layer (IPL). Double labeling with an antibody to choline acetyltransferase (ChAT) revealed that the GFP-band was located at strata 3 inner retina. The best-labeled GFP-expressing cell type in the INL was a wide-field amacrine cell that ramified in stratum 3. The GFP-expressing cells in the GCL resemble the type B1, or possibly A2 ganglion cells. The CD44-EGFP mice should provide a valuable resource for electrophysiological and connectivity studies of amacrine cells in the mouse retina.