[Lower urinary tract injury in transvaginal reconstructive pelvic surgery].

Objective: To explore the characteristics, prevention and treatment strategies of lower urinary tract injury in transvaginal reconstructive pelvic surgery (vRPS). Methods: A retrospective analysis was conducted on 24 patients who suffered lower urinary tract injuries occuring in vRPS from January 2005 to June 2021, among which 4 cases were referred to our hospital from other hospitals. Results: (1) In our hospital, 1 952 patients underwent vRPS for anterior and (or) middle pelvic organ prolapse during that study period, with a 1.0% (20/1 952) incidence of lower urinary tract injuries occurring in 20 cases. (2) Ureteral injuries were observed in 14 cases who underwent transvaginal high uterosacral ligament suspension (1.4%, 14/966). The symptoms were relieved after the removal of sutures. (3) Bladder injuries occurred in 6 cases in our hospital, with 4 cases (0.7%, 4/576) in anterior transvaginal mesh surgery (aTVM), one (0.4%, 1/260) in colpocleisis, and one (0.7%, 1/150) in apical suspension for fornix prolapse. An additional 4 cases of bladder injury were referred to our hospital after aTVM. Among the 8 cases of bladder injury during aTVM, 2 cases were intraoperative incidents. Cystoscopy confirmed that the superficial branch or puncture rod of anterior vaginal mesh had penetrated into the bladder. Re-puncturing and placement of the mesh were successfully performed. No abnormalities were observed during a follow-up period of 4-5 years. Postoperative bladder injuries were identified in 6 cases, characterized by mesh erosion into the bladder and formation of calculi. These injuries were confirmed between 6 months to 2 years after vRPS. The exposed mesh and calculi in the bladder were removed through laparotomy or cystoscopy, followed up for 2-12 years. One case experienced slight re-erosion of mesh to the bladder. Conclusions: Lower urinary tract injuries are difficult to avoid in vRPS, particularly in transvaginal high uterosacral ligament suspension and aTVM. However, the incidence is low. Lower urinary tract injuries during vRPS could be easily detected and managed intraoperatively because of the use of cystoscopy. As long-term postoperative complications, erosion of transvaginal mesh to lower urinary tract postoperatively could be treated correctly, seldom with severe sequelae.

[Effectiveness of abdominal minimal incision sacrocolpopexy for advanced pelvic organ prolapse].

Objective: To evaluate the indications, surgical skills and clinic outcomes of abdominal minimal incision sacrocolpopexy (AMISC) for treatment of advanced pelvic organ prolapse (POP). Methods: The retrospective study analyzed 30 women with advanced POP who underwent AMISC between June 2016 and October 2019, including 9 cases of recurrent prolapse and 10 cases of vault prolapse. AMISC was especially applicable to: (1) patients with several medical complications who was unable to tolerate general anesthesia or laparoscopic surgery, but able to tolerate combined spinal-epidural anesthesia and open surgery; (2) other abdominal procedures were indicated to perform with AMISC simultaneously, such as myomectomy, subtotal hysterectomy etc, the specimens were easy to get out of the abdominal cavity and morcellation was avoided; (3) surgeons preferring open surgery to laparoscopic surgery or skilled in open surgery; (4) patients with prior pelvic operations, presenting severe abdominal and pelvic adhesions. Objective outcomes were assessed by pelvic organ prolapse quantification (POP-Q) system. Subjective outcome were assessed by pelvic floor distress inventory-short form 20 (PFDI-20), pelvic floor impact questionnaire-short form (PFIQ-7) and patient global impression of improvement (PGI-I). Results: All patients with 1-3 medical complications were successfully performed with AMISC without stopping procedure, enlarging the incision or changing to other procedure, the operation duration was (110±19) minutes. The mean time of follow-up was (33.5±12.4) months (range: 8-49 months). The postoperative points of Aa, Ba, C, Ap, Bp reduced significantly and point C improved from (2.33±2.50) cm to (-7.54±1.18) cm after AMISC (P<0.01). The objective cure rates were both 100% (30/30) in apex and posterior compartment, while 97% (29/30) in anterior compartment. Postoperative scores of PFDI-20 and PFIQ-7 were all significant decreased (all P<0.01). About PGI-I, 29 patients chose "significant improvement", subjective satisfaction was 97% (29/30). Anterior sacral plexus hemorrhage occurred in 2 cases (7%, 2/30). There was no intestinal obstruction or injury of bladder, bowel and ureter intra- and postoperation. Two cases (7%, 2/30) had mesh exposure. Conclusion: AMISC is a safety, convenient, minimal traumatic and durable procedure for apical prolapse with short learning curve in the most of cases.

[Effect of ectodysplasin-A1 on proliferation and cell cycle of ameloblast-like cell].

Objective: To investigate the effects of ectodysplasin-A1 (EDA1) on the proliferation and cell cycle of ameloblast-like epithelial cells (LS8 cells). Methods: Wild EDA1 plasmid pCR3-Flag-EDA1-W (wild group), syndrome mutant EDA1 plasmid pCR3-Flag-EDA1-H252L (mutant group) and empty vector plasmid pCR3-Flag (control group) were transfected into LS8 cells. Cell proliferation was detected by methyl thiazolyl tetrazolium (MTT) assay and cell cycle was detected by flow cytometry. All tests were repeated three times. Results: Compared with the control group (0.105±0.032), the proliferation activity of the wild group (0.201±0.009) was significantly higher after 72 h (P<0.05). Compared with the control group (0.168±0.054) and the mutant group (0.194±0.059), the proliferation activity of the wild group (0.386±0.066) was significantly higher after 96 h (P<0.05). There was no significant difference between the mutant group and the control group at all time points (P>0.05). In the G0/G1 phase, compared with the control group (65.4%±2.1%) and the mutant group (66.6%±3.1%), the cell distribution ratio of the wild group (51.2%±1.1%) was significantly lower (P<0.01). In the S phase, compared with the control group (23.1%±2.0%) and the mutant group (21.9%±1.8%), the cell distribution ratio of the wild type group (37.3%±2.4%) was significantly higher (P<0.01). There was no significant difference in cell cycle distribution between the mutant group and the control group (P<0.05). Conclusions: Wild EDA1 promotes the proliferation of LS8 cells and the transformation from G0/G1 to S phase. The syndrome mutant EDA1 (EDA1-H252L) loses its function of regulating the cell proliferation and cell cycle of LS8 cells.

Modulation of rumen fermentation and microbial community through increasing dietary cation-anion difference in Chinese Holstein dairy cows under heat stress conditions.

AIMS: The effect of increasing dietary cation-anion difference (DCAD) on rumen fermentation and ruminal microbial community in dairy cows under heat stress (HS) conditions were evaluated. METHODS AND RESULTS: This study was performed as a two-period cross-over design during the summer season, with eight lactating dairy cows randomly distributed to either a control DCAD diet (CON: 33·5 mEq/100 g DM) or high DCAD diet (HDCAD: 50·8 mEq/100 g DM). Throughout the present study, the temperature and humidity index (THI; 80·2 ± 4·29) was generally elevated above the threshold (THI = 72) that is reported to cause HS in lactating dairy cows. Rumen liquid samples were collected on 15 and 21 d during each 21 d-period. The absolute concentration of ruminal total volatile fatty acid (TVFA) in HDCAD treatment was significantly (P < 0·05) higher than those in the control, whilst the ruminal pH, NH3 -N, and VFA molar percentages were unaffected through increasing DCAD. Furthermore, the copy numbers of the cellulolytic bacteria Ruminococcus albus and Ruminococcus flavefaciens in rumen fluid significantly (P < 0·05) rose along with the increment of DCAD. Although the Alpha diversity indexes and the bacterial microbiota structure were unaffected, increasing DCAD significantly (P < 0·05) enriched the phylum Fibrobacteres and genus Fibrobacter in the microflora of rumen fluid, whilst the genera Flexilinea and Dubosiella were the most differentially abundant taxa in the control. CONCLUSIONS: Increasing DCAD under HS conditions resulted in a greater concentration of total VFA without affecting rumen bacteria diversity or structure, although the enrichment of some cellulolytic/hemicellulolytic bacteria was observed. SIGNIFICANCE AND IMPACT OF THE STUDY: The present study provides information on the modulation of rumen fermentation and microbial community through the increment of DCAD in Holstein dairy cows under HS conditions.

[A five-year analysis of effect on transvaginal high uterosacral ligament suspension with or without native-tissue repair for middle compartment defect].

Objective: To assess the five-year effect of the transvaginal high uterosacral ligament suspension (HUS) with or without additional concomitant native-tissue anterior and (or) posterior repair in women suffering from middle compartment defect. Methods: A retrospective review of records identified 79 women who underwent transvaginal HUS with or without additional concomitant native-tissue anterior and (or) posterior repair from January 2007 to January 2018 in Fourth Medical Center, General Hospital of People's Liberation Army. The middle compartment defects were predominant in these patients with point C no less than point Ba or Bp if accompanied with anterior or posterior vaginal wall prolapse. Follow-up visits were performed 2,6 and 12 months after surgery and then annually. Anatomic results of pelvic organ prolapse (POP) was established by pelvic examination using pelvic organ prolapse quantitation system (POP-Q) staging. Funtional results were obtained by patient global impression of improvement (PGI-I) scale in POP, pelvic floor distress inventory-short form 20 (PFDI-20) and pelvic floor impact questionnaire short form (PFIQ-7). Surgical success required the fulfillment of all 3 criteria: (1) anterior or posterior vaginal wall prolapsed leading edge of 0 cm or less and apex of 1/2 total vaginal length or less; (2) the absence of POP symptoms as reported on the PFDI-20 question No.3 ( "Do you usually have a bulge or something falling out that you can see or feel in your vaginal area?" ); and (3) no prolapse reoperations or pessary use during the study period. Results: Of 79 women, 51(65%, 51/79) women completed the five-year follow-up during the study period. The median follow-up time was 5.2 years (2.8-8.3 years). The overall surgery success rate was 86% (44/51) according to above all 3 criteria. Prolapse recurrence rates were isolated anterior 8% (4/51), isolated apical 0, isolated posterior 2% (1/51) and multiple compartments 4% (2/51). Seven women (14%,7/51) developed anterior or posterior prolapse beyond the hymen with the leading edge≤1 cm. No apical prolapsed occurred. None of recurrent women underwent retreatment,including either surgery or pessary usage at last follow-up. The subjective satisfaction rate was 90% (46/51). There was a 1% (1/79) rate of intraoperative ureteral kinking and 3% (2/79) rate of postoperative morbidity. Conclusions: The transvaginal HUS for middle compartment defect offers good long-term anatomical results with excellent vault suspension. With additional concomitant native-tissue anterior and (or) posterior repair, it will be a reconstructive surgery for the majority of moderate-to-severe POP. It is minimal traumatic and worthy of being popularized for clinical application.

[Effectiveness of vaginal high uterosacral ligament suspension for treatment of recurrent pelvic organ prolapse].

Objective: To evaluate the indications and clinic outcomes of vaginal high uterosacral ligament suspension (HUS) for treatment of recurrent advanced pelvic organ prolapse (POP). Methods: This retrospective study analyzed 42 women with recurrent advanced POP who were referred to Fourth Medical Center of PLA General Hospital and underwent transvaginal HUS between November 2005 and January 2018. Primary surgeries included 30 vaginal colporrhaphy, 5 Manchester operation, 5 transvaginal mesh repair,2 sacrospinous ligament fixation.The median time for recurrence from primary pelvic floor repair surgery was 9 months, including 14 cases (33%, 14/42) ≤3 months (median time was 2 months) and 25 cases (67%, 28/42) longer than 3 months (median time was 18 months).The rate of recurrent prolapse in stage Ⅲ or Ⅳ was 79% (33 cases), 45% (19 cases) and 17%(7 cases) in anterior, apical and posterior compartment respectively. Results: Transvaginal high bilateral uterosacral ligaments were identified and used for successful vaginal vault suspension after vaginal hysterectomy and residual cervical resection in all 42 consecutive patients. The cases of transvaginal mesh used in anterior wall and posterior wall were 25 (60%, 25/42) and 3 (7%, 3/42) respectively. There was no major intra- and postoperative complications,such as ureter and other pelvic organ injury. The median time of follow-up was 5.3 years after transvaginal HUS. The points of pelvic organ prolapse quantification system reduced significantly and point C improved from +0.3 cm to -8.2 cm after reoperation (P<0.01). The objective cure rate were 100% (42/42) both in apex and posterior compartment,while 93% (39/42) in anterior compartment. None had reoperation or pessary usage for recurrence of prolapse. Conclusion: Transvaginal HUS with vaginal wall repair could be as a safety, cost-effective, minimal traumatic and durable procedure for recurrent POP in the most of cases.

Transcriptome analysis of osteoblasts in an ovariectomized mouse model in response to physical exercise.

OBJECTIVES: Osteoporosis is a metabolic disease resulting in progressive loss of bone mass as measured by bone mineral density (BMD). Physical exercise has a positive effect on increasing or maintaining BMD in postmenopausal women. The contribution of exercise to the regulation of osteogenesis in osteoblasts remains unclear. We therefore investigated the effect of exercise on osteoblasts in ovariectomized mice. METHODS: We compared the activity of differentially expressed genes of osteoblasts in ovariectomized mice that undertook exercise (OVX+T) with those that did not (OVX), using microarray and bioinformatics. RESULTS: Many inflammatory pathways were significantly downregulated in the osteoblasts after exercise. Meanwhile, IBSP and SLc13A5 gene expressions were upregulated in the OVX+T group. Furthermore, in in vitro assay, IBSP and SLc13A5 mRNAs were also upregulated during the osteogenic differentiation of MC3T3-E1 and 7F2 cells. CONCLUSION: These findings suggest that exercise may not only reduce the inflammatory environment in ovariectomized mice, indirectly suppressing the overactivated osteoclasts, but may also directly activate osteogenesis-related genes in osteoblasts. Exercise may thus prevent the bone loss caused by oestrogen deficiency through mediating the imbalance between the bone resorptive activity of osteoclasts and the bone formation activity of osteoblasts.Cite this article: W-B. Hsu, W-H. Hsu, J-S. Hung, W-J. Shen, R. W-W. Hsu. Transcriptome analysis of osteoblasts in an ovariectomized mouse model in response to physical exercise. Bone Joint Res 2018;7:601-608. DOI: 10.1302/2046-3758.711.BJR-2018-0075.R2.

[Clinical management of 110 cases of polypropylene mesh and sling exposure after reconstructive pelvic floor surgery].

Objective: To explore the clinical management and outcomes of polypropylene mesh and sling exposure after reconstructive pelvic surgery (RPS) . Methods: A total of 110 cases of mesh and sling exposure after RPS were analyzed, who admitted between Jan. 2002 and Oct. 2017 in First Affiliated Hospital of PLA General Hospital, in which 3 cases were referred from other hospitals. Mesh and sling exposures were identified in the outpatient clinic and categorized and managed according to International Continence Society and International Urogynecology Association (ICS-IUGA) classification about category, time and site (CTS) of mesh complication. Outpatient management included observation, topical estrogen use and mesh removal. Management in hospital included surgical removal of exposed mesh and repair of the resulting defects under the anesthesia. Seventy-four cases were managed in the outpatient setting, and 36 cases required inpatient management. Follow-up was consecutively performed from 1 month to 10 years. Objective outcome included the surgeon's assessment of the healing state of the vaginal mucosa. Subjective outcome was evaluated with patient global impression of improvement questionnaire (PGI-I) . Results: One hundred and ten patients with mesh exposure were classified according to the different RPS underwent. There were 95 cases from transvaginal mesh surgery, 5 cases from anti-stress urinary incontinence sling surgery, and 10 cases from sacrocolpopxy. The outpatient group healed at an average of (3.0±1.8) months. Of the 36 patients who required inpatient management, 21 cases healed completely at an average of 7 days after one surgery. The remaining 8 cases required either two or three times surgeries or conservative management. In the outpatient group, the PGI-I scale very much better was found in 65 cases (87.8%) and much better in 9 cases (12.2%) . In the inpatient surgery group, the scale was very much better in 30 cases (83.3%) , and much better in 6 cases (16.7%) . Conclusions: Among patients with mesh exposure after mesh-augmented RPS, 2/3 of patients with a CTS classification 1-3 could be managed in the office, and remaining 1/3 with CTS classification 4-6 need operation under anesthesia in hospital. If the mesh and sling exposure could be scientifically classified, according to the size, site and accompany symptoms, as well as pain, most of the mesh complications after explosure could be resolved. Using the pelvic floor repair and polypropylene mesh sling, the majority of the patients could get a better outcome, without affecting the effect of the original operation.

Author Correction: WNT-activated bone grafts repair osteonecrotic lesions in aged animals.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

WNT-activated bone grafts repair osteonecrotic lesions in aged animals.

The Wnt pathway is a new target in bone therapeutic space. WNT proteins are potent stem cell activators and pro-osteogenic agents. Here, we gained insights into the molecular and cellular mechanisms responsible for liposome-reconstituted recombinant human WNT3A protein (L-WNT3A) efficacy to treat osteonecrotic defects. Skeletal injuries were coupled with cryoablation to create non-healing osteonecrotic defects in the diaphysis of the murine long bones. To replicate clinical therapy, osteonecrotic defects were treated with autologous bone graft, which were simulated by using bone graft material from syngeneic ACTB-eGFP-expressing mice. Control osteonecrotic defects received autografts alone; test sites received autografts treated ex vivo with L-WNT3A. In vivo µCT monitored healing over time and immunohistochemistry were used to track the fate of donor cells and assess their capacity to repair osteonecrotic defects according to age and WNT activation status. Collectively, analyses demonstrated that cells from the autograft directly contributed to repair of an osteonecrotic lesion, but this contribution diminished as the age of the donor increased. Pre-treating autografts from aged animals with L-WNT3A restored osteogenic capacity to autografts back to levels observed in autografts from young animals. A WNT therapeutic approach may therefore have utility in the treatment of osteonecrosis, especially in aged patients.

[Long-term effectiveness of transvaginal high uterosacral ligament suspension].

Objective: To assess the long-term effectiveness of the transvaginal high uterosacral ligament suspension (HUS) in women suffering from advanced pelvic organ prolapse (POP). Methods: A retrospective review of records identified 118 women who underwent transvaginal HUS with or without additional concomitant anterior and (or) posterior repairs from June 2003 to August 2009 in the First Affiliated Hospital, General Hospital of People's Liberation Army. Of 118 women, 104 women completed the follow-up during study period; these 104 women were analysed. Follow-up visits were performed 2, 6 and 12 months after surgery and then annually. Anatomic results of POP was established by pelvic examination using pelvic organ prolapse quantitation system (POP-Q) staging. Funtional results were obtained by patient global impression of improvement (PGI-I), pelvic floor distress inventory-short form 20 (PFDI-20) and pelvic floor impact questionnaire short form (PFIQ-7). Surgical success required the fulfillment of all 3 criteria: (1) prolapse leading edge of 0 cm or less and apex of 1/2 total vaginal length or less; (2) the absence of pelvic organ prolapse symptoms as reported on the PFDI-20 question No. 3 ( "Do you usually have a bulge or something falling out that you can see or feel in your vaginal area?" ); and (3) no prolapse reoperations or pessary use during the study period. Results: The mean follow-up time was (9.1±1.5) years. The overall surgery success rate was 91.3% (95/104) according to above all 3 criteria. Prolapse recurrence rates were isolated anterior 6.7% (7/104), isolated apical 0, isolated posterior 2.9% (3/104) and multiple compartments 1.0% (1/104). Five women (4.8%, 5/104) developed bothersome vaginal bulge symptoms. None of recurrent women underwent retreatment, including either surgery or use of a pessary at last follow-up. The subjective satisfaction rate was 90.4% (94/104). PFDI-20 and PFIQ-7 scores showed a statistically significant improvement from preoperative 72 and 65 points to postoperative 17 and 9 points respectively (all P<0.01). There was a 2.9% (3/104) rate of intraoperative ureteral kinking and 3.8% (4/104) rate of postoperative morbidity. Conclusions: The transvaginal HUS for vault prolapse offers good long-term anatomical results with excellent vault suspension. With additional concomitant anterior and (or) posterior repairs, it will be a reconstructive surgery for the majority of advanced POP. It is minimal traumatic and appropriate for different type of POP, especially for the eldly patients. It is worthy of being popularized for clinical application.

[Follow-up of survived children supported by extracorporeal membrane oxygenation].

Objective: To summarize the follow-up of children treated with venoarterial extracorporeal membrane oxygenation (ECMO) by incision of internal carotid artery-vein. Method: This was a retrospective study, 10 children in whom the conventional therapy failed, but survived after treatment with ECMO technique through right internal jugular artery-venous incision and ligation after ECMO in pediatric intensive care unit (PICU) of Fudan University were followed up from December 2011 to December 2015. Primary disease situation, neurological development, growth and development, quality of life and personality development were included. All children were followed up once every six months after discharge, and once a year after 2 years. The longest follow-up period was 2 years. Result: Of the 10 children 6 were boys and 4 girls, and ranged in age of onset from 1 day to 12 years . Five children had neurological problems, including abnormal brain CT (n=3), abnormal electroence phalogram (EEG) (n=3), and mental retardation (n=2). Three of those children had a history of hypoxia, 2 of whom accepted cardiopulmonary resuscitation. Eight children had normal brainstem evoked potential (BEAP) examinations. Primary diseases were all well controlled except for 6 children who had different degrees of sequelae, including mild arrhythmia (n=1), heart structure abnormality (n=2) and abnormal pulmonary function (n=3). The sequelae caused by the primary diseases were just embodied in examination. Their head circumference measurement and physical examination showed no obvious abnormalities, and blood biochemical tests, hemoglobin and blood gas analysis were also normal. One child's height was less than P3. The one who accepted cardiopulmonary resuscitation (CPR) at early stage of primary disease had trouble in daily life, presenting uncoordinated and unstable walking. One child was slightly bad tempered and asocial, and the rest were normal. Conclusion: The rest could take care of themselves. The ECMO sequelae of those survived children were mostly caused by primary diseases. ECMO treatment is safe and has less adverse consequences.

[Complications of critically ill children supported by venoarterial extracorporeal membrane oxygenation].

OBJECTIVE: To study the complications occurred in the process of venoarterial extracorporeal membrane oxygenation (VA-EMCO) in critically ill children. METHOD: To analyze retrospectively 25 children who were admitted to PICU of Children's Hospital of Fudan University from December 2011 to December 2015. They were all treated with VA-ECMO with incision and catheterization in right internal carotid artery and jugular vein. Complications were recorded during ECMO process. RESULT: The duration of ECMO treatment was 14 to 567 h (153 (112, 204) h). Seventeen cases (68%) withdrew from ECMO successfully and 15 cases (60%) survived to discharge. Thirty-three complications occurred during ECMO treatment. Of which, mechanical complications occurred 9 times, including oxygenator leakage (n=3), hemolysis (n=2), water tank failure (n=2), pump head rupture (n=1) and piping thrombosis (n=1). Somatic complications appeared 24 times, including neurological complications (n=8) which included cerebral infarction (n=2), convulsions (n=2), intracranial hemorrhage (n=2), thrombosis after ligation of internal jugular vein (n=1) and cerebral atrophy (n=1); bleeding complications (n=8) which included bleeding at puncture sites (n=4), ECMO canalized site bleeding (n=3), and spontaneous (n=1), intracranial hemorrhage or hematoma cases(n=2) in mechanical complications; acute kidney injury (n=5) and microcirculation thrombosis (n=3). CONCLUSION: ECMO technique may cause complications, which mainly include neurological, mechanical and clotting complications. Advanced equipments and materials and well-managed teamwork are helpful in decreasing complications.

De novo symptoms and their impact on life quality in patients following transvaginal reconstructive pelvic surgery with polypropylene mesh.

OBJECTIVE: To study the de novo symptoms and their impact on life quality in patients that underwent transvaginal reconstructive pelvic surgery (RPS) with polypropylene mesh. MATERIALS AND METHODS: From May 2004 to March 2011, 114 severe pelvic organ prolapse (POP) patients with Stage III-IV by POP-Q system underwent RPS with polypropylene mesh. Patients completed pelvic floor distress inventory short form (PFDI-20) and pelvic floor impact questionnaire short form (PFIQ-7) preoperatively and repeated them at two and six months, and one year postoperatively. RESULTS: Ninety-six (84%, 96/114), 85 (75%, 85/114), and 77 (68%, 77/114) patients, respectively, completed the questionnaires at two months, six months, and one year postoperatively. All patients had POP-Q staging scoring < or = I at one year after surgery. Nineteen (19.8%, 19/96) patients had mesh exposure at two and six months (7.8%, 6/77) at one year follow-up. Most vaginal and pelvic symptoms, urinary and obstructive defecation bothersome symptoms improved significantly at two months postoperatively and this improvement was maintained at the one year follow-up. Mean score of PFDI-20 and PFIQ-7 all improved significantly postoperatively at two and six months and at one year follow-up (p < 0.01). Fifty percent (48/96) of patients had postoperative de novo symptoms at the two months follow-up predominantly presented with bothersome vaginal discharge (35.4%, 34/96) and pelvic muscle symptoms (20.8%, 20/96). Patients with de novo symptoms had higher postoperative mean POPDI-6 and POPIQ-7 scores (p < 0.05) than those without at the two month follow-up, but no significant difference was seen at the six month and at one year follow-ups. Patients with bothersome vaginal discharge had higher vaginal mesh exposure rate (41.2%, 14/34) than patients without (8.1%, 5/62) (p = 0.0003). One year after operation, 77 (68%) patients completed the non-validated satisfaction questionnaire. Seventy-four (96%, 74/77) patients said that they were either 'very satisfied' or 'satisfied' with the outcome of their surgery, while three (4%, 3/77) reported unsatisfactory results. CONCLUSIONS: De novo symptoms were common after transvaginal RPS with polypropylene mesh, but most of them were moderate and resolved within six months postoperatively and seldomly had a long-term negative impact on their quality of life. The impact of dyspareunia on patients' sexual function requires further research.

Evidence for protein-mediated fatty acid efflux by adipocytes.

AIM: The hormonally controlled mobilization and release of fatty acids from adipocytes into the circulation is an important physiological process required for energy homeostasis. While uptake of fatty acids by adipocytes has been suggested to be predominantly protein-mediated, it is unclear whether the efflux of fatty acids also requires membrane proteins. METHODS: We used fluorescent fatty acid efflux assays and colorimetric assays for free fatty acids and glycerol to identify inhibitors with effects on fatty acid efflux, but not lipolysis, in 3T3-L1 adipocytes. We assessed the effect of these inhibitors on a fibroblast-based cell line expressing fatty acid transport protein 1, hormone-sensitive lipase and perilipin, which presumably lacks adipocyte-specific proteins for fatty acid efflux. RESULTS: We identified 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) as an inhibitor of fatty acid efflux that did not impair lipolysis or the cellular exit of glycerol but lead to an accumulation of intracellular fatty acids. In contrast, fatty acid efflux by the reconstituted cellular model for fatty acid efflux was responsive to lipolytic stimuli, but insensitive to DIDS inhibition. CONCLUSION: We propose that adipocytes specifically express an as yet unidentified DIDS-sensitive protein that enhances the efflux of fatty acids and therefore may lead to novel treatment approaches for obesity-related disorders characterized by abnormal lipid fluxes and ectopic triglyceride accumulation.

Stimulation of lipolysis and hormone-sensitive lipase via the extracellular signal-regulated kinase pathway.

Hormonally stimulated lipolysis occurs by activation of cyclic AMP-dependent protein kinase (PKA) which phosphorylates hormone-sensitive lipase (HSL) and increases adipocyte lipolysis. Evidence suggests that catecholamines not only can activate PKA, but also the mitogen-activated protein kinase pathway and extracellular signal-regulated kinase (ERK). We now demonstrate that two different inhibitors of MEK, the upstream activator of ERK, block catecholamine- and beta(3)-stimulated lipolysis by approximately 30%. Furthermore, treatment of adipocytes with dioctanoylglycerol, which activates ERK, increases lipolysis, although MEK inhibitors decrease dioctanoylglycerol-stimulated activation of lipolysis. Using a tamoxifen regulatable Raf system expressed in 3T3-L1 preadipocytes, exposure to tamoxifen causes a 14-fold activation of ERK within 15-30 min and results in approximately 2-fold increase in HSL activity. In addition, when differentiated 3T3-L1 cells expressing the regulatable Raf were exposed to tamoxifen, a 2-fold increase in lipolysis is observed. HSL is a substrate of activated ERK and site-directed mutagenesis of putative ERK consensus phosphorylation sites in HSL identified Ser(600) as the site phosphorylated by active ERK. When S600A HSL was expressed in 3T3-L1 cells expressing the regulatable Raf, tamoxifen treatment fails to increase its activity. Thus, activation of the ERK pathway appears to be able to regulate adipocyte lipolysis by phosphorylating HSL on Ser(600) and increasing the activity of HSL.

Characterization of the functional interaction of adipocyte lipid-binding protein with hormone-sensitive lipase.

Hormone-sensitive lipase (HSL) is an intracellular lipase that plays an important role in the hydrolysis of triacylglycerol in adipose tissue. HSL has been shown to interact with adipocyte lipid-binding protein (ALBP), a member of the family of intracellular lipid-binding proteins that bind fatty acids and other hydrophobic ligands. The current studies have addressed the functional significance of the association and mapped the site of interaction between HSL and ALBP. Incubation of homogeneous ALBP with purified, recombinant HSL in vitro resulted in a 2-fold increase in substrate hydrolysis. Moreover, the ability of oleate to inhibit HSL hydrolytic activity was attenuated by co-incubation with ALBP. Co-transfection of Chinese hamster ovary cells with HSL and ALBP resulted in greater hydrolytic activity than transfection of cells with HSL and vector alone. Deletional mutations of HSL localized the region of HSL that interacts with ALBP to amino acids 192-200, and site-directed mutagenesis of individual amino acids in this region identified His-194 and Glu-199 as critical for mediating the interaction of HSL with ALBP. Interestingly, HSL mutants H194L and E199A, each of which retained normal basal hydrolytic activity, failed to display an increase in hydrolytic activity when co-transfected with wild type ALBP. Therefore, ALBP increases the hydrolytic activity of HSL through its ability to bind and sequester fatty acids and via specific protein-protein interaction. Thus, HSL and ALBP constitute a functionally important lipolytic complex.

Absence of cardiac lipid accumulation in transgenic mice with heart-specific HSL overexpression.

Hormone-sensitive lipase (HSL) hydrolyzes triglyceride (TG) in adipose tissue. HSL is also expressed in heart. To explore the actions of cardiac HSL, heart-specific, tetracycline (Tc)-controlled HSL-overexpressing mice were generated. Tc-responsive element-HSL transgenic (Tg) mice were generated and crossed with myosin heavy chain (MHC)alpha-tTA Tg mice, which express the Tc-responsive transactivator (tTA) in the heart. The double-Tg mice (MHC-HSL) were maintained with doxycycline (Dox) to suppress Tg HSL. Upon removal of Dox, cardiac HSL activity and protein increased 12- and 8-fold, respectively, and the expression was heart specific. Although cardiac TG content increased twofold in control mice after an overnight fast, it did not increase in HSL-induced mice. Electron microscopy showed numerous lipid droplets in the myocardium of fasted control mice, whereas fasted HSL-induced mice showed virtually no droplets. Microarray analysis showed altered expression of cardiac genes for fatty acid oxidation, transcription factors, signaling molecules, cytoskeletal proteins, and histocompatibility antigens in HSL-induced mice. Thus cardiac HSL plays a role in controlling accumulation of triglyceride droplets and can affect the expression of a number of cardiac genes.

Human BMP-7/OP-1 induces the growth and differentiation of adipocytes and osteoblasts in bone marrow stromal cell cultures.

We studied the effects of BMP-7/OP-1 on growth and differentiation of bone marrow stromal cells. BMS2, a mouse bone marrow stromal cell line capable of differentiating into adipocytes and osteoblasts, were treated in a serum-free medium containing differentiation agents that favor the expression of both lineages. BMP-7/OP-1 stimulated cell proliferation and differentiation concomitantly. These effects were dose- and growth phase-dependent. Cells were more sensitive to the treatment early in the culture (30-40% confluence) with a significant increase in cell proliferation and markers of differentiation at low concentrations. When treated later in the growth phase (90-100% confluence), no significant increase in cell proliferation was seen. The concentration requirement for cells later in the culture to reach an equivalent degree of differentiation was 3-10- fold higher than for cells treated early. In both cases, the effects on adipocyte differentiation were biphasic; low concentrations stimulated adipocyte differentiation which was inhibited at higher concentrations where stimulation of osteoblast markers were observed. We conclude that cell proliferation and cell differentiation into adipocyte/osteoblast can occur simultaneously under BMP-7/OP-1 treatment.