Evaluation of Autophagy Process in Differentiation of Human Induced Pluripotent Stem Cells toward Insulin Producing Cells.

Background: Autophagy is an intracellular self-degradative homeostasis process which eliminates undesirable and harmful macromolecules and organelles. Autophagy is also involved in self-renewal and differentiation of induced pluripotent stem cell (iPSCs). Objective: In this study, we investigated the expression profile of autophagy marker genes in human iPSCs during their differentiation induction toward insulin producing ß-like cells. Methods: Human iPSC line, R1-hiPSC1, was used for differentiation induction toward ß-like cells. The mRNA expression of Nanog, OCT4 (pluripotency markers), SOX17, FOXA2 (endodermic markers), PTF1A, NKX6.1 (exocrine/endocrine determinants), and PDX1 were measured during differentiation stages. Autophagy was monitored by genes expression study of four autophagy markers, MAP1LC3B, BECN1, SQSTM1/P62 and ATG5, along with protein expression profile of LC3b-II during differentiation stages. Results: The mRNA expression measurement of pluripotency, endoderm and exocrine/endocrine marker genes confirmed that hiPSCs skipped pluripotency, differentiated into endoderm, passed through the pancreatic lineage commitment stage and successfully generated insulin producing ß-like cells. Expression profile of autophagy genes during differentiation stages indicated the decreased expression levels at the early stages (EB and MEI) and then increased at the definitive endoderm stages (DEI 1, DEI 2 and DE) followed by a subtractive pattern toward the end of differentiation. The results of protein expression of LC3b-II were consistent with gene expression data. Conclusion: This study demonstrated the high contribution of key autophagy genes/proteins during the differentiation of hiPSC toward ß-like cells. The enhanced autophagy levels were a prominent feature of early stages of differentiation and DE rather than the later stages.

Predicting changes in the status of patient-reported outcome measures after Birmingham Hip Resurfacing: an observational cohort study with a median follow-up of ten years.

AIMS: It is not known whether change in patient-reported outcome measures (PROMs) over time can be predicted by factors present at surgery, or early follow-up. The aim of this study was to identify factors associated with changes in PROM status between two-year evaluation and medium-term follow-up. PATIENTS AND METHODS: Patients undergoing Birmingham Hip Resurfacing completed the Veteran's Rand 36 (VR-36), modified Harris Hip Score (mHHS), Tegner Activity Score, and Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC) at two years and a minimum of three years. A change in score was assessed against minimal clinically important difference (MCID) and patient-acceptable symptom state (PASS) thresholds. Binary logistic regression was used to assess the relationship between patient factors and deterioration in PASS status between follow-ups. RESULTS: Overall, 18% of patients reported reductions in mHHS total score exceeding MCID, and 21% reported similar reductions for WOMAC function scores. Nonetheless, almost all patients remained above PASS thresholds for WOMAC function (98%) and mHHS (93%). Overall, 66% of patients with mHHS scores < PASS at two years reported scores > PASS at latest follow-up. Conversely, 6% of patients deteriorated from > PASS to < PASS between follow-ups. Multivariable modelling indicated body mass index (BMI) > 27 kg/m2, VR-36 Physical Component Score (PCS) < 51, VR-36 Mental Component Score (MCS) > 55, mHHS < 84 at two years, female sex, and bone graft use predicted these deteriorating patients with 79% accuracy and an area under the curve (AUC) of 0.84. CONCLUSION: Due to largely acceptable results at a later follow-up, extensive monitoring of multiple PROMs is not recommended for Birmingham Hip Resurfacing patients unless they report borderline or unacceptable hip function at two years, are female, are overweight, or received a bone graft during surgery. Cite this article: Bone Joint J 2019;101-B:1431-1437.

The outcome and survival of metal-on-metal hip resurfacing in patients aged less than 50 years: a prospective observational cohort study with minimum ten-year follow-up.

AIMS: The aim of this study was to report the implant survival and patient-reported outcome measures (PROMs) in a consecutive series of patients aged less than 50 years at the time of arthroplasty using the Birmingham Hip Resurfacing system (BHR), with a minimum follow-up of ten years. PATIENTS AND METHODS: A total of 226 patients with osteoarthritis of the hip, who underwent BHR and presented to a single surgeon, were included in the study. Survival of the implant was confirmed by cross-checking with the Australian Orthopaedic Association National Joint Replacement Registry. Kaplan-Meier survival curves with 95% confidence intervals (CIs) were constructed. Pre- and postoperative PROMs were compared with t-tests, and postoperative scores were compared using anchor analysis with age and gender matched normative data. RESULTS: At median follow-up of 12 years (interquartile range (IQR) 10 to 13), six BHRs were revised, with a cumulative rate of survival of 96.8% (95% confidence interval (CI) 94.2 to 99.4) at 15 years, and with a significantly lower (p = 0.019) cumulative rate of revision than the national average for the same device at ten years. Most revisions (n = 4) were undertaken early, less than three years postoperatively, and occurred in women. Patient-reported general health (Veteran's Rand-36), disease state (Western Ontario and McMaster Universities Osteoarthritis Index), function (modified Harris Hip Score) and level of activity (Tegner activity score) maintained significant (p < 0.01 for each) improvements beyond ten years postoperatively and were equal to, or exceeded, age- and gender-matched normative data in more than 80% of the patients. CONCLUSION: Longer term PROMs after BHR, from a single surgeon, for patients aged less than 50 years remain under-reported. We found that the outcome after a BHR, at a minimum of ten years postoperatively, remained satisfactory, particularly for self-reported hip function.

What happens to femoral neck bone mineral density after hip resurfacing surgery?

The major advantage of hip resurfacing is the decreased amount of bone resection compared with a standard total hip replacement. Fracture of the femoral neck is the most common early complication and poor bone quality is a major risk factor. We undertook a prospective consecutive case control study examining the effect of bone mineral density changes in patients undergoing hip resurfacing surgery. A total of 423 patients were recruited with a mean age of 54 years (24 to 87). Recruitment for this study was dependent on pre-operative bilateral femoral bone mineral density results not being osteoporotic. The operated and non-operated hips were assessed. Bone mineral density studies were repeated over a two-year period. The results showed no significant deterioration in the bone mineral density in the superolateral region in the femoral neck, during that period. These findings were in the presence of a markedly increased level of physical activity, as measured by the short-form 36 health survey physical function score.

Primary and long-term stability of a short-stem hip implant.

The new generation short-stem hip implants are designed to encourage physiological-like loading, to minimize stress-strain shielding and therefore implant loosening in the long term. As yet there are no long-term clinical studies available to prove the benefits of these short-stem implants. Owing to this lack of clinical data, numerical simulation may be used as a predictor of longer term behaviour. This finite element study predicted both the primary stability and long-term stability of a short-stem implant. The primary implant stability was evaluated in terms of interface micromotion. This study found primary stability to fall within the critical threshold for osseointegration to occur. Longer term stability was evaluated using a strain-adaptive bone remodelling algorithm to predict the long-term behaviour of the bone in terms of bone mineral density (BMD) changes. No BMD loss was observed in the classical Gruen zones 1 and 7 and bone remodelling patterns were comparable with hip resurfacing results in the literature.

Nocardia nova septic arthritis following total knee replacement: a case report.

We describe a case of Nocardia nova septic arthritis following a total knee replacement. A 55-year-old obese woman was admitted to hospital 5 months after knee surgery with a 3-week history of pain, swelling, and restricted mobility in her left knee but no preceding trauma/injury. 30 ml of cloudy joint fluid was aspirated and an arthroscopic examination showed extensive fibrin formation and synovitis. An arthroscopic washout was carried out using 16 litres of saline, followed by total synovectomy and intensive antibiotic therapy (clarithromycin 500 mg twice daily and co-trimoxazole [sulphamethoxazole 400 mg and trimethoprim 80 mg] once daily and augmentin duo forte 875 mg twice daily). At 2.5 years, the patient had recovered completely with no prosthetic loosening.

Periprosthetic bone remodelling of a collum femoris preserving cementless titanium femoral hip replacement.

Total hip arthroplasty represents a major surgical achievement for pain relief and restoration of lifestyle quality due to the joint disease of osteoarthritis. Total hip replacement has evolved over the past 30 years utilising a variety of biocompatible materials, geometric shapes and fixation techniques. The main objective of this study is to investigate the long-term effects of strain adaptive bone remodelling due to the influence of a novel titanium cementless femoral hip replacement. The period of on-growth has been taken into account and the simulation has been run to predict the remodelling behaviour for a 36-month period. The main conclusion from this analysis is that the implant does shield the calcar to a similar degree as other cementless femoral hip designs. It does, however, tend to cause bone to be laid down along its length. This may, in part, be due to the novel geometry of the implant interlocking with and loading the bone.

Vertebral morphometry reference data by X-ray absorptiometry (MXA) in Iranian women.

The anterior, medial, and posterior heights and the A/P and M/P ratios of the spine (T5-L4) in 41 normal premenopausal Iranian women were determined using an imaging densitometer (Expert XL) and dual energy X-ray absorptiometry (DXA) method. All the women were healthy (age 20-39 years, and height 149-171 cm), without any signs of vertebral fractures, and with normal bone mineral density (BMD) of the spine and femoral neck (T-score>-1.5). The vertebral heights were normalized using the Expert XL software, and the average vertebral height for the L2-L4 vertebrae was taken to minimize the effect of variation of body size among the subjects. The Z-score for all vertebral heights (T5-L4) averaged -0.68, with the A/P and M/P ratios coming to +0.34 and +0.49, respectively. It showed the normalization procedure not to correct the differences of vertebral heights in Iranian women. The average of the three heights (H (avg)) correlated fairly well with the stature of the subject (r=0.47, p<0.05), but no correlation was found between H (avg) and subject age (p>0.05). The lower vertebral heights in older women in comparison with the younger women (0.4 mm) obtained in our study can be attributed to the relatively shorter stature of older women (mean 154 vs 159 cm for younger women, p<0.05). It was concluded that the normalization procedure used in the software does not equally apply to Iranian women due to their having different heights than those of American and northern European women, from whom the reference data for the Expert XL software have been gathered. The reference values thus obtained are therefore not accurate for our population group and a separate study with a bigger and more varied sample group is needed for obtaining more definitive results.

Surgical management of aneurismal dilation of vein and pseudoaneurysm complicating hemodialysis arteriovenuos fistula.

BACKGROUND: Pseudoaneurysm (PS) and aneurismal dilation of vein (ADV) are recognized complications of arteriovenous fistulas (AVF) in patients on hemodialysis. We present our experience about surgical management of these complications, which resulted in AVF preservation for continuing hemodialysis. MATERIAL AND METHODS: Twenty-two patients underwent surgical repair of an aneurismal dilation of vein or a pseudoaneurysm arising from a native AVF. In 14 patients the aneurismal dilation of vein arose from the venous limb of AVF and in eight patients the pseudoaneurysm arose from an arteriovenous anastomotic site in the antecubital and anterior part of arm. The mean follow-up period was 15 months. Clamp Aneurysm Repair (CAR) was performed to repair the aneurismal dilation of venous limb of AVF and Tourniquet Aneurysm Repair (TAR) was performed to repair PS that arose from AVF in the antecubital and anterior part of arm. RESULTS: In eight of the 14 patients with aneurismal dilation of vein, who underwent CAR procedure, vascular access was preserved. In three patients with aneurismal dilation of vein in snuffbox and one in forearm, the AVF had failed due to prior venous thrombosis of AVF. In two of 14 patients, there was no need for preservation of AVF because of renal transplantation. The technical success rate and patency rate during follow up period in CAR method was 100%. In seven of eight patients with psudoaneurysm in the antecubital and anterior part of arm, who underwent TAR procedure, the AVF remained patent. The technical success rate in TAR method was 87.5%, and the patency rate was 87.5%. Overall, technical success rate was 95.45% and patency rate was 93.75%. During the 15 months of follow up period hemodialysis program through the repaired AVF sustained as desired. CONCLUSIONS: The surgical methods used in our study could effectively repair the aneurismal dilation of vein and psudoaneurysm arising from a native AVF, and it lead to preservation of the AVF patency for continuing hemodialysis. These methods are technically feasible, safe and cost-effective procedures. It does not require dissection and additional incision for control of the vein and artery proximal and distal to the aneurismal dilation of vein and pseudoaneurysm; result in shorter time of procedure without complications.

Sterilization of contaminated bone-tendon autografts using 10% povidone-iodine solution.

This study evaluates methods of sterilizing contaminated bone-tendon autografts using 10% povidone-iodine solution. Sterile grafts were prepared from human cadavers. Grafts were immersed in a suspension of either Staphylococcus aureus or Pseudomonas aeruginosa, and three sets of sterilization experiments were performed in 10% povidone-iodine for 30 minutes: one each with S. aureus and P. aeruginosa by static soaking and a third with S. aureus by serial washing with agitation. Of grafts inoculated with S. aureus, six of six grew the test organism after soaking at room temperature, as did five of six after soaking at 36 degrees C and also eight of nine after washing with agitation. Of grafts inoculated with P. aeruginosa, five of six grew the test strain after soaking at room temperature, as did six of six after soaking at 36 degrees C. Thirty minutes of exposure to aqueous 10% povidone-iodine does not adequately sterilize an inoculated graft.

The cold plate as a test of nociceptive behaviors: description and application to the study of chronic neuropathic and inflammatory pain models.

A cold plate apparatus was designed to test the responses of unrestrained rats to low temperature stimulation of the plantar aspect of the paw. At plate temperatures of 10 degrees C and 5 degrees C, rats with either chronic constriction injury (CCI) of the sciatic nerve or complete Freund's adjuvant (CFA) induced inflammation of the hindpaw displayed a stereotyped behavior. Brisk lifts of the treated hindpaw were recorded, while no evidence of other nociceptive behaviors could be discerned. The most consistent responses were obtained with a plate temperature of 5 degrees C in three 5-min testing periods, separated by 10-min intervals during which the animals were returned to a normal environment. Concomitantly to cold testing, the rats were evaluated for their response to heat (plantar test) and mechanical (von Frey hairs) stimuli. In both injury models, while responses to heat stimuli had normalized at 60 days post-injury, a clear lateralization of responses to cold was observed throughout the entire study period. Systemic lidocaine, clonidine, and morphine suppressed responses to cold in a dose-related fashion. At doses that did not affect motor or sensory behavior, both lidocaine and its quaternary derivative QX-314 similarly reduced paw lifts, suggesting that cold hyperalgesia is in part due to peripheral altered nociceptive processing. Clonidine was more potent in CCI then in CFA rats in reducing the response to cold. Paradoxically, clonidine increased the withdrawal latencies to heat in the CCI hindpaw at 40 days and thereafter, at a time when both hindpaws had the same withdrawal latencies in control animals. Morphine was also more potent on CCI than CFA cold responses, indicating that, chronically, CFA-induced hyperalgesia might be opiate resistant. Evidence for tonic endogenous inhibition of cold hyperalgesia was obtained for CFA rats, when systemic naltrexone significantly increased the number of paw lifts; this was not found in rats with CCI. At 60 days, neither morphine nor naltrexone affected cold-induced paw lifting in CFA rats, suggesting that the neuronal circuit mediating cold hyperalgesia in these animals had become opiate insensitive. In conclusion, the cold plate was found to be a reliable method for detecting abnormal nociceptive behavior even at long intervals after nerve or inflammatory injuries, when responses to other nociceptive stimuli have returned to near normal. The results of pharmacological studies suggest that cold hyperalgesia is in part a consequence of altered sensory processing in the periphery, and that it can be independently modulated by opiate and adrenergic systems.

Cytokines and matrix metalloproteinases mRNA expression in archival human tissues from failed total hip arthroplasty using in situ hybridization and color video image analysis.

Reports on the histological and biochemical nature of periprosthetic fibrous/granulomatous tissue has, to date, been largely limited to frozen tissue sections. This study reports the cytokine and matrix metalloproteinase profiles found in periprosthetic interface tissues in THA which have failed due to aseptic loosening and in capsular tissues obtained at primary surgery. The study employs immunohistochemistry, in situ hybridization, and color video image analysis on formalin fixed and paraffin embedded sections.

Imaging of the liver with helical CT: a survey of scanning techniques.

OBJECTIVE: We conducted a survey of the members of the Society of Computed Body Tomography/Magnetic Resonance to assess current techniques in liver imaging using helical CT. MATERIALS AND METHODS: The survey, which was designed to update earlier surveys from 1987 and 1993, included a questionnaire distributed to 77 members of the Society of Computed Body Tomography/Magnetic Resonance. RESULTS: Forty-nine members responded, representing 28 institutions. In 1993, 19% (5/26) of institutions used helical scanners, compared with 82% (23/28) in 1996. The group of institutions with helical CT served as the focus of this survey. In 1993, 58% of institutions used 1-cm collimation: in 1996, 78% (18/23) used thinner, 7- to 8-mm collimation. In 1987, 41% used power injectors compared with 85% in 1993 and 100% in 1996. In 1996, monophasic injections were used by 96% (22/23) of institutions. In 1993, most institutions used a contrast material injection rate of 1.5-2.0 ml/sec; in 1996, most used a 2.5-3.0 ml/sec injection rate. In 1993, 96% of institutions used 125-150 ml of contrast material; in 1996, 57% (13/23) of institutions used 125-150 ml and 30% of institutions used less than 125 ml of contrast material. A delay time of 21-45 see was used by 83% of institutions in 1993, whereas in 1996, 83% (19/23) of institutions used a longer delay time of 50-80 sec. In 1996, 13% of institutions used an individual scan delay technology and all institutions performed multiphasic scanning of hypervascular lesions. CONCLUSION: The availability of helical CT has changed radiologists' approach to liver imaging. The greatest effects of which are a more widespread use of power injectors, longer delay times, thinner collimation, increased contrast material injection rates, decreased contrast material volumes, and multiphasic scanning.

SPECT for acute knee pain.

In the assessment of acute knee pain following recent trauma frequently related to sporting activities, bone scintigraphy was performed on 52 patients; in 40 of these patients, scintigraphy immediately preceded arthroscopy. SPECT was critical in providing delineation and sitting of the abnormalities; this would not have been feasible with conventional planar imaging in 45 of the patients. Multiple sites of focal uptake were frequently visualized at ligamentous insertions or in intra-articular areas of cartilaginous erosion. Damaged anterior cruciate ligaments were only identified in 5 out of 10 patients, occurring when there was avulsion of the tibial attachment, but changes resulting from patellar injury were demonstrated in all 8 patients. The most characteristic scintigraphic change resulted from meniscal tears, which was diagnosed in 31 out of 35 patients in whom the lesion had been suspected clinically. In the identification of this common injury, SPECT had a sensitivity of 88%, specificity of 87%, and diagnostic accuracy of 88%. Thus, although trauma readily induces scintigraphic abnormalities in and around the knee, the patterns of alteration associated with particular lesions can be identified by SPECT and can provide considerable assistance in management, particularly in determining the need for arthroscopy.

Adult osteosclerosis.

Quantitative bone histology was carried out in five osteosclerotic adults. The bone was extremely hard in all patients, and open biopsy was usually required. One patient, aged 18 years, presented with hypoplastic anemia, and the most probable explanation for the osteosclerosis is a marrow stem cell defect leading to defective osteoclasts. Another had the dominant form of osteopetrosis. Her bone contained cartilage remnants, and there were many large, morphologically abnormal osteoclasts, which lacked normal cytoplasmic acid phosphatase activity. The third patient had chronic renal failure and osteomalacia; here the increased bone mass might have resulted from an inability of normal osteoclasts to resorb bone, due to the surface coating of osteoid, though an earlier increase of bone formation cannot be excluded. The fourth patient, who suffered from systemic mastocytosis, had high turnover bone, with greatly increased bone formation. The fifth patient, with fluorosis of bone, also had increased bone formation and resorption, the process being much more pronounced in the head of her pathologically fractured femur than it was in the iliac crest. In this patient some osteoclasts had reduced acid phosphatase activity and long cytoplasmic extensions, both changes similar to those observed in diphosphonate-treated animals. Very diverse processes can result in the increased cancellous bone mass producing the radiographic appearance of diffuse osteosclerosis.

Femoral shaft fractures in children: the effect of initial shortening on subsequent limb overgrowth.

Twenty-six children between the ages of ten months and 15 years with closed fractures of the femoral shaft treated by closed means were reviewed to determine whether shortening at the time of fracture union overcame the problem of subsequent limb overgrowth. The mean shortening at the time of union was 9.8 millimeters. The mean overgrowth was 8.8 mm. The initial shortening and subsequent limb overgrowth were related, with a correlation coefficient of 0.63 (p less than 0.01). The greater the initial shortening, the stronger appeared to be the stimulus for overgrowth.