Application of open porous poly(D,L-lactide-co-glycolide) microspheres and the strategy of hydrophobic seeding in hepatic tissue cultivation.

In this article, porous poly(D,L-lactide-co-glycolide) (PLGA) microsphere scaffolds with a size of ∼ 400 µm and pores of ∼ 20 µm were prepared for constructing injectable three-dimensional hepatocyte spheroids. The porous sites of PLGA microspheres provided a spatial space for hepatocyte distribution. Hepatocytes spheroids were cocultured with human umbilical vein endothelial cell, bone marrow mesenchymal stem cell, or NIH/3T3 cells by combining the porous PLGA microspheres with the relatively hydrophobic culture strategy. The combination of open porous microspheres, hepatocytes, and nonparenchymal cells was demonstrated for application in functional hepatic tissue reconstruction. Hepatocellular-specific functions can sustained up to 2 weeks in the support of coculturing with nonparenchymal cells. The spheroidal hepatocyte coculture system had the advantages of an injectable delivery, higher cell seeding density, protection from exerted shear stress, better exchange of nutrients, oxygen and metabolites, and heterotypic cell-cell contact within and between microspheres.

Clinical outcomes and development of children born after intracytoplasmic sperm injection (ICSI) using extracted testicular sperm or ejaculated extreme severe oligo-astheno-teratozoospermia sperm: a comparative study.

OBJECTIVE: To evaluate the clinical outcomes and development of children born after intracytoplasmic sperm injection (ICSI) with extracted testicular sperm or ejaculated extreme severe oligo-astheno-teratozoospermia (OAT) sperm. DESIGN: Retrospective study. SETTING: Infertility clinic at Chang Gung Memorial Hospital. PATIENT(S): A total of 126 ICSI cycles were performed using extracted testicular sperm from men with azoospermia and 65 ICSI cycles using fresh ejaculated sperm from men with extreme severe OAT. INTERVENTION(S): Retrospective analysis of clinical outcomes and development of children born after ICSI with extracted testicular sperm or ejaculated extreme severe OAT sperm. MAIN OUTCOME MEASURE(S): Fertilization rates, number of grade 1 zygotes and number of embryos produced, implantation rate, clinical pregnancy rate, abortion and live birth rate per transfer, perinatal outcomes, and birth defects. RESULT(S): The demographic and clinical factors, including age, E(2) level on hCG day, number of oocytes retrieved, normal fertilization rate, zygote grade 1 score distribution, number of top-quality embryos transferred, clinical pregnancy rate per transfer, chemical pregnancy rate per transfer, implantation rate, live birth rate per transfer, and abortion rate per transfer, were similar between the groups. Sixty live births resulted from 48 extracted testicular sperm cycles and 21 live births from 19 extreme severe OAT. The obstetric and perinatal outcomes were similar between the groups, and children conceived by using ICSI were healthy and without major psychomotor or intellectual development retardation. One case of tetralogy of Fallot occurred in each group. CONCLUSION(S): There is no evidence of differences in the clinical outcomes and development of children result after ICSI with extracted testicular sperm or ejaculated extreme severe OAT sperm.

High-efficiency cell seeding method by relatively hydrophobic culture strategy.

Cell adhesion efficiency is one of the key factors affecting the results of manufacturing tissue engineering constructs. High efficiency is required for seeding low proliferation cells onto scaffolds. In this study, we designed a strategy to improve the efficiency of cell adhesion using hydrophobic cell culture environment to enhance cells adhering to a scaffold. Cells have lower affinity to the surface of polydimethylsiloxane (PDMS) than tissue culture polystyrene (TCPS) plates. When cells were cultured with gelatin microspheres or chitosan films in a PDMS-coated plate instead of a normal TCPS plate, there was a significant increase in cell attachment efficiency. Cells cultured in the PDMS-coated system tended to selectively attach onto the gelatin microspheres or chitosan films, which are relatively more hydrophilic than the PDMS surface. However, minimal cell attachment on gelatin microspheres or chitosan films was observed when gelatin microspheres or chitosan films were placed in normal TCPS plate. Cell counting experiments with gelatin microspheres in the PDMS-coated system resulted in a cell attachment efficiency of 89.8% after 1 day of cultivation, whereas the cell attachment efficiency was less than 1% in normal TCPS plate. The results demonstrate that the method is easy to use and could be useful for fast cultivation of cell-scaffold constructs.

Large-scale cultivation of transplantable dermal papilla cellular aggregates using microfabricated PDMS arrays.

In this work we have developed a strategy for cultivating dermal papilla (DP) cells to form multiple arrayed spheroidal microtissues for transplantation on a micropatterned polydimethylsiloxane (PDMS)-based tissue culture polystyrene (TCPS) plate system. We also describe the behavior of dermal papilla cells on this platform and the spontaneous formation of spheroidal microtissues by DP cells. We used a hydrophobic PDMS arrayed chip as a master to separate the seeded cells in the TCPS culture plate. By controlling the cell seeding densities, a microwell with arrayed DP spheroidal microtissues was easily formed. Formation of DP microtissues was associated with overlapping multilayered cells on the microwells and low cell-substrate adhesivity on the PDMS film. The microwell environment enhanced the aggregation of DP cells into spheroidal microtissues on the TCPS culture plate. The spheroidal microtissues preserved their hair induction potential in vitro and in vivo. A large quantity of DP spheroidal microtissues could be obtained rapidly and simply using this platform. We could harvest hundreds of DP microtissues (352 microtissues) with a cell seeding density of 1×106 cells well⁻¹ after 3 days cultivation in one well of a 24-well plate. This is the first demonstration of the formation of DP spheres in large quantitites.

Patterned PDMS based cell array system: a novel method for fast cell array fabrication.

Cell-cell interaction is important in numerous biological processes, including cell growth, cell differentiation and migration. The ability to generate pre-determined cell patterns or cell arrays on a study surface is crucial for cell-cell interaction studies. In this paper, we developed a method for fast cell array fabrication using laser sintering and the hydrophobicity of PDMS films. This approach can be easily adopted and is cost-effective. Hydrophobic PDMS films were fabricated into polymeric chips containing hundreds of microwells. The films were then transferred onto tissue culture surfaces to separate cells in the formation of cell arrays (Patterned PDMS based cell array system, PCAS). We used NIH/3T3 fibroblast cells to demonstrate the feasibility of PCAS. The success of fast fabrication of patterned cell arrays was obtained using different initial cell seeding densities. We also used poorly adherent PC-12 cells to demonstrate the cell-cell communication. Results showed that the method is very useful for studying topics such as cell-cell interaction, cell-substrate interaction or cell migration.

The role of the preoperative serum carcinoembryonic antigen level in early-stage adenocarcinoma of the uterine cervix.

OBJECTIVE: The purpose of the study was to identify the relationship between preoperative serum levels of carcinoembryonic antigen (CEA) and clinicopathological variables in early-stage adenocarcinoma of the uterine cervix. METHODS: From February 1990 to August 2002, 117 patients with surgically treated early-stage cervical adenocarcinoma that had had preoperative serum CEA evaluations were retrospectively reviewed. The cut-off value for CEA, based on the manufacturer's recommendations, was 5 ng/ml. For an evaluation of the relationship between the clinicopathological factors and increased levels of serum tumor markers, the Chi-Square/Fisher's exact test and logistic regression were used for univariate and multivariate analysis, respectively. RESULTS: The mean age of the patients was 46 years (range, 21-78). Of the 117 patients, 28 had preoperative serum CEA levels greater than 5 ng/ml. In a univariate analysis, the increased marker was associated with a larger tumor size, presence of lymphovascular invasion, and deeper cervical wall invasion. However, in a multivariate analysis, the preoperative CEA level had a significant impact on the determination of the depth of stromal invasion (OR 4.12, 95% CI 1.97-8.68, P < 0.001). CONCLUSION: In early-stage cervical adenocarcinoma, preoperative serum CEA levels seem to be useful in estimating the depth of cervical stromal invasion. Assessment of tumor antigen CEA levels should be integrated with the routine examination in the work-up of patients with adenocarcinoma of the uterine cervix.

Multiple uterine angioleiomyomas in a woman presenting with severe menorrhagia.

BACKGROUND: Angioleiomyoma is a rare benign neoplasm that originates from smooth muscle cells and contains thick-walled vessels. There were only five cases of uterine angioleiomyoma reported in the available English literatures. We present here the unique computed tomography finding in a patient with multiple uterine angioleiomyomas causing severe menorrhagia. CASE: A 50-year-old, nulligravid woman consulted us with the complaint of menorrhagia for 3 years and progressively palpable lower abdominal mass for a half year. Laboratory findings were all within normal limits except lower hemoglobin concentration (6.2 g/dl). An abdomino-pelvic computed tomography (CT) showed that a huge 30-cm heterogeneously multilobulate mass with solid and laminated configuration, with cystic and multiseptal contents was found in left lower abdomen and pelvic cavity. At laparotomy, the area beneath the left broad ligament was filled with a well-encapsulated, elastic, ovoid, and lobulate mass that connected to the uterus and measured up to 20 cm in greatest diameter. The uterus was composed of a huge intramural tumor and measured 28 x 21 x 12 cm. The uterus and huge subserosal tumor were resected completely and a frozen section was obtained. The final histopathologic diagnosis was angioleiomyoma, a definitely benign soft tissue tumor. Eighteen months after surgery there was no recurrence. CONCLUSION: Uterine angioleiomyoma should be considered when prominent tortuous vascular-like enhancing structures are noted on CT examination of a well-demarcated soft tissue mass arising from the uterus in pelvis. Either angiomyomectomy with tumor-free margins or hysterectomy proved to be an effective treatment in these cases, and resulted in a good recovery and a satisfactory outcome.

Major complications associated with laparoscopic-assisted vaginal hysterectomy: ten-year experience.

STUDY OBJECTIVE: To describe our experience with major complications associated with laparoscopic-assisted vaginal hysterectomy (LAVH) and compare our results with those of the American Association of Gynecologic Laparoscopists (AAGL) membership survey and another similar study. DESIGN: Retrospective study (Canadian Task Force classification II-3). SETTING: University-affiliated hospital. PATIENTS: Two thousand seven hundred two women. Intervention. LAVH. MEASUREMENTS AND MAIN RESULTS: Demographic data and medical histories (age, parity, surgical indications, pathologic findings, major complications) were analyzed. Major complications were 11 bladder injuries, 4 ureter injuries, 11 bowel injuries, 2 vascular injuries, 2 cases of massive bleeding from the vaginal cuff or colpotomy wound with associated impending shock, 2 cases of postoperative ileus, and 2 pelvic abscesses. Our overall major complication rate was 1.3% compared with 2.7% in the AAGL 1995 membership survey (p <0.001). Similar rates of febrile morbidity (2.2% and 2.0%), bleeding requiring transfusion (0.05% and 0.06%), and bowel, ureteral, or bladder injury (1.0% and 1.0%) were noted between our study and the other 1995 study (all p >0.05). Of 34 major complications in our study, 24 occurred during hysterectomy performed by inexperienced general gynecologists and 10 by an experienced endoscopist (p = 0.005). CONCLUSION: The rate of major complications associated with LAVH can be reduced when the procedure is performed by a well-trained laparoscopic surgeon compared with a less-experienced general gynecologist.

The predictive value of using a combined Z-score and day 3 embryo morphology score in the assessment of embryo survival on day 5.

BACKGROUND: The purpose of this study was to evaluate the ability of using the Z-score alone, or, in combination with the day 3 embryo morphology score, to predict embryo viability at day 5 from a large cohort of embryos derived from patients undergoing treatment with IVF/ICSI. METHODS AND RESULTS: In this retrospective study, a total of 1894 zygotes from 346 treatment cycles (295 couples) was analysed between January 2001 and May 2002. The Z-scoring system was useful in predicting day 5 embryo survival. The mean +/- SD day 5 embryo survival rates were 78.2 +/- 1.7, 49.0 +/- 2.5, 21.4 +/- 3.2 and 11.8 +/- 5.6% for Z-1, Z-2, Z-3 and Z-4 zygotes groups respectively. Embryos derived from Z-1 scores and grade I day 3 embryo scores showed the best day 5 embryo survival and a very high implantation potential. CONCLUSIONS: These data suggest that a combined evaluation of the Z-score and day 3 embryo morphology is highly predictive of embryo outcome after IVF/ICSI. The Z-score could be of great help in the selection of embryos for cultures extended to later stages. The Z-score alone, or preferably in combination with day 3 embryo morphology, is useful in the determination of the most suitable embryos and the number of embryos for transfer, thus achieving the optimal chance of conception while reducing the risk of high order multiple pregnancy.

Leiomyosarcoma of the uterus: a clinicopathologic study of 21 cases.

BACKGROUND: The purpose of the present study was to identify the prognostic factors of and to determine the most appropriate mode of treatment for uterine leiomyosarcoma. METHODS: We reviewed the hospital records, including surgical notes and pathologic reports, of 21 patients with uterine leiomyosarcoma treated at Chang Gung Memorial Hospital, Kaohsiung, Taiwan, between 1987 and 1997. Univariate analysis was performed using the log-rank test. Cox regression was used to identify independent prognostic factors. RESULTS: The mean follow-up time was 30 months, and the 5-year disease-free survival rate was 55%. Evaluating the correlation between clinicopathologic parameters and survival, early stage (p = 0.0002), tumor cells without necrosis (p = 0.0026), low-grade tumor (p = 0.015), absence of vascular space involvement (p = 0.006), and tumor without atypia (p = 0.016) were associated with good prognosis. However, in a multivariate analysis using the Cox model, only advanced stage (p = 0.032) and tumor necrosis (p = 0.032) were found to be independent poor prognostic factors. The 5-year disease-free survival was only 21% in patients with the presence of any one or both of these two factors. Five of 11 patients in this group had received aggressive adjuvant therapy after surgery, and none of them survived more than 19 months. CONCLUSIONS: We found that patients with advanced stage or presence of tumor necrosis had an extremely poor prognosis. Adjuvant therapy seemed to play a limited role, and provided no survival benefit. Treatment for these patients should be palliative until effective therapeutic modalities prove otherwise.

Can a preoperative CA 125 level be a criterion for full pelvic lymphadenectomy in surgical staging of endometrial cancer?

OBJECTIVE: The aim of the present study was to evaluate whether a preoperative serum level of CA 125 in patients with endometrial cancer can provide additional information in determining the extent of lymphadenectomy required in the surgical staging and which cutoff value is optimal in this respect. METHODS: CA 125 was measured in 124 patients diagnosed with endometrial carcinoma who underwent surgery at our institution between January 1995 and May 2000. Statistic analysis was performed using the Mann-Whitney U test and Kruskal-Wallis test to evaluate the association of preoperative CA 125 levels with various factors. The chi(2)/Fisher's exact test and a logistic regression model were employed to examine the effects of clinicopathological factors on serum CA 125 levels. Furthermore, a receiver operating characteristic curve was used to determine which cutoff value of the preoperative CA 125 was the optimal one. RESULTS: Univariate analyses showed that elevated CA 125 levels were significantly correlated with an advanced stage, larger tumor size, increasing depth of the myometrial invasion, cervical invasion, positive cytology, and lymph node metastases (P < 0.05). Multivariate analyses using a logistic regression model showed lymph node metastases had the most significant effect on the elevation of CA 125 levels. The ROC curve determined that the best cutoff value was 40 U/ml; the sensitivity and specificity for screening lymph node metastases were found to be 77.8 and 81.0%, respectively. CONCLUSION: Our data provide evidence indicating that a preoperative CA 125 level greater than 40 U/ml can be considered a criterion for full pelvic lymphadenectomy in the surgical staging of endometrial cancer.