A potential osteogenic role for microRNA-181a-5p during palatogenesis.

BACKGROUND: In a previous study, we found that the highly conserved hsa-miR-181a-5p is downregulated in palatal fibroblasts of non-syndromic cleft palate-only infants. OBJECTIVES: To analyze the spatiotemporal expression pattern of mmu-miR-181a-5p during palatogenesis and identify possible mRNA targets and their involved molecular pathways. MATERIAL AND METHODS: The expression of mmu-miR-181a-5p was analyzed in the developing palates of mouse embryos from E11 to E18 using qPCR and ISH. Mouse embryonic palatal mesenchyme cells from E13 were used to analyze mmu-miR-181a-5p expression during osteogenic differentiation. Differential mRNA expression and target identification were analyzed using whole transcriptome RNA sequencing after transfection with a mmu-miR-181a-5p mimic. Differentially expressed genes were linked with underlying pathways using gene set enrichment analysis. RESULTS: The expression of mmm-miR-181a-5p in the palatal shelves increased from E15 and overlapped with palatal osteogenesis. During early osteogenic differentiation, mmu-miR-181a-5p was upregulated. Transient overexpression resulted in 49 upregulated mRNAs and 108 downregulated mRNAs (adjusted P-value < 0.05 and fold change > ± 1.2). Ossification (Stc1, Mmp13) and cell-cycle-related GO terms were significantly enriched for upregulated mRNAs. Analysis of possible mRNA targets indicated significant enrichment of Hippo signaling (Ywhag, Amot, Frmd6 and Serpine1) and GO terms related to cell migration and angiogenesis. LIMITATIONS: Transient overexpression of mmu-miR-181a-5p in mouse embryonic palatal mesenchyme cells limited its analysis to early osteogenesis. CONCLUSION: Mmu-miR-181-5p expression is increased in the developing palatal shelves in areas of bone formation and targets regulators of the Hippo signaling pathway.

Excess vitamin a might contribute to submucous clefting by inhibiting WNT-mediated bone formation.

OBJECTIVES: Cleft lip and/or palate (CLP) is a common craniofacial birth defect caused by genetic as well as environmental factors. The phenotypic spectrum of CLP also includes submucous clefts with a defect in the palatal bone. To elucidate the contribution of vitamin A, we evaluated the effects of the vitamin A metabolite all-trans retinoic acid (ATRA) on the osteogenic differentiation and mineralization of mouse embryonic palatal mesenchymal cells (MEPM). SETTING AND SAMPLE POPULATION: MEPM cells were isolated from the prefusion palates of E13 mouse embryos from three different litters. MATERIALS AND METHODS: MEPM cells were cultured with and without 0.5 µM ATRA in osteogenic medium. Differentiation was analysed by the expression of osteogenic marker genes and alkaline phosphatase (ALP) activity after 1, 2, and 7 days. The expression of Wnt marker genes was also analysed. Mineralization was assessed by alizarin red staining after 7, 14, 21, and 28 days. RESULTS: The bone marker genes Sp7, Runx2, Alpl, and Col1a1 were inhibited 10% ± 2%, 59% ± 7%, 79% ± 12% and 57% ± 20% (P < .05) at day 7. ALP activity was inhibited at days 1 and 7 by 35 ± 0% (P < .05) and 23 ± 6% (P < .001). ATRA also inhibited mineralization at 3 and 4 weeks. Finally, expression of the universal Wnt marker gene Axin2 was strongly reduced, by 31 ± 18% (P < .001), at day 7. CONCLUSION: Our data indicate that ATRA (vitamin A) inhibits bone formation by reducing Wnt signalling. This might contribute to the molecular aetiology of submucous clefting.

Assessing clinical trunk change with surface topography: anterior scoliosis correction as a model to evaluate curve progression.

PURPOSE: Previous work has suggested that surface topography can be used for repeated measurements of deformity during curve monitoring following an initial radiograph. Changes in deformity during natural curve progression may be subtle. An important preemptive question to answer is whether topography can follow a large change in spine deformity, as in scoliosis correction. We assess the ability of surface topography to track the evolution of spine deformity during anterior scoliosis correction relative to traditional radiographs. Anterior scoliosis correction was chosen for this analysis because it changes the shape of the trunk without leaving a surgical scar and muscle atrophy along the posterior spine. METHODS: Following IRB approval, 18 patients aged 14.6 ± 2.0 years at surgery were enrolled in a retrospective review of coronal radiographs and topographic scans acquired before and after scoliosis correction. Radiographic and topographic measures for the coronal curve angle before and after surgery were compared. RESULTS: Surface topography estimates correlate with radiographic measures of the pre- (r = 0.7890, CI = [0.4989 0.9201], p < 0.00001), postsurgical (r = 0.7485, CI = [0.4329 0.9006], p = 0.0004), and the change in the coronal curve angle (r = 0.6744, CI = [0.3028 0.8680], p = 0.0021) due to surgery. CONCLUSIONS: We provide evidence open for further extension that topography can follow changes in the coronal curve angle comparably to radiographs. LEVEL OF EVIDENCE: Level IV.

Progressive double major scoliotic curve with concurrent lumbosacral spondylolisthesis in a skeletally immature patient with Marfan syndrome treated with anterior scoliosis correction.

STUDY DESIGN: Case report (review of patient records, imaging, and pulmonary function tests) and literature review. OBJECTIVES: To describe the case of a skeletally immature patient with Marfan syndrome who underwent anterior scoliosis correction (ASC) and muscle-sparing posterior far lateral interbody fusion (FLIF) in a two-stage procedure to correct progressive severe double major scoliosis and spondylolisthesis. Patients with Marfan syndrome suffer from rapidly progressive scoliosis and spondylolisthesis. Operative treatment has typically been limited to PSF, but newer techniques may be less invasive and provide more spine motion. METHODS: A 12-year-old girl with Marfan syndrome, spondylolisthesis, and severe progressive scoliosis underwent a two-stage procedure to achieve correction. Muscle-sparing posterior FLIF of the spondylolisthesis from L4-S1 was initially performed, followed 1 week later by ASC from right T4-T11 and left T11-L3 using an anterior screw/cord construct. RESULTS: Follow-up from the index procedures for the spondylolisthesis and scoliosis is 35 months. No significant complications occurred in perioperative and postoperative follow-up periods. At the 13-month follow-up, the double major scoliosis showed continued curve correction via growth modulation and overcorrection of the lumbar to - 13°. A revision lengthening procedure of the anterior cord from T11-L3 was performed. An asymptomatic elevated hemidiaphragm was discovered at 6 weeks postoperation, which was believed to be secondary to retraction neuropraxia and subsequently improved. At 21 months postlengthening and 35 months postindex procedure, she is skeletally mature and the curves have maintained correction in both the coronal and sagittal planes without any further complications. CONCLUSIONS: Anterior scoliosis correction of both a thoracic and lumbar curve combined with an L4-S1 PSF was effective for this patient and may be promising for patients with Marfan syndrome, progressive scoliosis, and spondylolisthesis. Overcorrection can be planned for and easily corrected by inserting a new cord of a different length.

Pediatric Spine Disorders.

Pediatric spine disorders are numerous and are quite different when compared with the adult population. This article focuses on some of the more common pediatric spine disorders. This article summarizes such disorders and discusses typical treatment options in the pediatric orthopedic armamentarium.

Retinoic acid disrupts osteogenesis in pre-osteoblasts by down-regulating WNT signaling.

The skull bones are formed by osteoblasts by intramembranous ossification. WNT signaling is a regulator of bone formation. Retinoic Acid (RA) act as a teratogen affecting craniofacial development. We evaluated the effects of RA on the differentiation and mineralization of MC-3T3 cells, and on the expression of WNT components. MC-3T3 were cultured with or without 0.5 µM RA in osteogenic medium and mineralization was assessed by alizarin red staining. The expression of osteogenic marker genes and WNT genes was evaluated at several time points up to 28 days. RA significantly inhibited MC-3T3 mineralization (p < 0.01), without affecting ALP activity or Alp gene expression. Both parameters gradually increased in time. During culture, RA stimulated Runx2 expression at 14 and 28 days compared to the respective controls (p < 0.05). Also, RA significantly reduced Sp7 expression at days 14 and 21 (p < 0.05). Simultaneously, RA significantly reduced the expression of the WNT genes cMyc, Lef1, Lrp5, Lrp6 and Wnt11 compared to the controls (p < 0.05). In contrast, RA increased the expression of the WNT inhibitors Dkk1 at day 21 and Dkk2 at days 14 and 21 (p < 0.01). Our data indicate that RA disrupts osteogenic differentiation and mineralization by inhibiting WNT signaling.

Progressive Neuromuscular Scoliosis Secondary to Spinal Cord Injury in a Young Patient Treated With Nonfusion Anterior Scoliosis Correction.

Ninety-eight percent of skeletally immature patients with spinal cord injury (SCI) suffer from progressive neuromuscular scoliosis (NMS). Operative treatment has typically been limited to posterior spinal fusion (PSF), but a newer technique as described may be less invasive and preserve more function. A PSF of the entire spine to the pelvis is standard of care. However, maintenance of spinal flexibility, motion, and potential growth is desirable. We present a case for proof-of-concept of utilizing a surgical motion-preserving technique to treat progressive NMS in an 11year-old girl with T10 level (AIS B) paraplegia with a progressive 60° NMS of the lumbar spine. She had anterior scoliosis correction (ASC) from T11-L5 without fusion. Over 24 months, the curve growth-modulated to a residual of 12° with continued modulation to 7° at 3-year follow-up (skeletal maturity).

Diagnostic value of early postoperative MRI and diffusion-weighted imaging following trans-sphenoidal resection of non-functioning pituitary macroadenomas.

AIM: To establish the value of early contrast-enhanced magnetic resonance imaging (MRI) and diffusion-weighted imaging (DWI) in differentiating residual pituitary adenoma from postoperative surgical changes. MATERIALS AND METHODS: Thirty patients with non-functioning pituitary macroadenomas, who were undergoing trans-sphenoidal adenomectomy, were prospectively studied. Patients were imaged with both MRI and DWI in the early postoperative period, as well as 6-months post-surgery. Patterns of postoperative contrast enhancement were described (non-enhancement, peripheral enhancement, and nodular enhancement). Apparent diffusion coefficient (ADC) maps were utilised to select the region of interest (ROI) for ADC calculations. RESULTS: Seventeen patients had postoperative surgical granulation tissue and 13 had residual adenoma based on the 6 months follow-up imaging. Mean ADC values of postoperative granulation tissue and residual adenoma were 1.476±0.476×10-3 mm2/s and 0.855±0.190×10-3 mm2/s, respectively, in the early postoperative period, and 1.357±0.416×10-3 mm2/s and 0.829±0.201×10-3 mm2/s, respectively, at the 6-month follow-up. ADC values of granulation tissue were significantly different from that of residual adenoma at both time points (p<0.001). Sensitivity, specificity, positive and negative predictive values of early MRI were 84.6%, 94.1%, 91.7%, and 88.9% respectively, and of early DWI were 91%, 97%, 94.3%, and 93%, respectively. CONCLUSION: Early postoperative DWI after trans-sphenoidal resection of pituitary macroadenomas may be more helpful than early MRI in differentiating residual adenoma from post-surgical changes.

Anesthetic considerations for a novel anterior surgical approach to pediatric scoliosis correction.

BACKGROUND: Idiopathic scoliosis is a condition that may require surgical correction. Limitations of previous surgical modalities, however, created the need for novel methods of repair. One such technique, a newer form of anterolateral scoliosis correction, has shown considerable promise, which our center has had substantial experience performing. AIM: In this article, we present the case details of our first 105 patients for the purposes of describing the evolution and details of the anesthetic management and considerations for this procedure. METHODS: A retrospective review of medical records for 105 patients undergoing anterolateral instrumentation procedure for idiopathic scoliosis correction done at a single institution from May 2014 to June 2016 was performed. The details of perioperative management as well as surgical technique were reported for all patients. RESULTS: The mean age for patients was 14.8 years (range 10-18); the mean weight was 49.9 kg (range 25-82). Unilateral procedures were performed on 46.7%, with bilateral and hybrid procedures performed on 50.5% and 4.7%, respectively. The median number of levels corrected was 8 (interquartile range [IQR] 7-9) for unilateral, right 7 (IQR 6-7) and left 5 (IQR 4-5) for bilateral, and 4 (IQR 4-4.5) for hybrids. The average estimated blood loss (EBL) was 310 mL±138, with cell salvaged blood transfused in 61% of patients, and allogenic blood transfusion required in only two patients. CONCLUSIONS: The described anesthetic and analgesic management provides a framework for delivering perioperative care for this challenging procedure, which is gaining popularity as a modality for scoliosis correction.

Validation of a lactic acid- and citric acid-based antimicrobial product for the reduction of Escherichia coli O157: H7 and Salmonella on beef tips and whole chicken carcasses.

The objectives of this study were to determine the effects of a lactic acid- and citric acid-based antimicrobial product on the reduction of Salmonella on whole broiler carcasses during processing and the reduction of Salmonella and Escherichia coli O157:H7 on beef trim. Freshly harvested broiler carcasses were inoculated with an inoculum of Salmonella strains to yield a 10(5) CFU/ml pathogen load on the surface of the carcass. The beef tips were inoculated as well with an inoculum of either E. coli O157:H7 or Salmonella to yield 10(4) CFU/100 cm(2). After 30 min for attachment, the broiler carcasses were treated with Chicxide applied for 5 s via a spray or immersed in Chicxide for 5, 10, or 20 s. Broiler carcasses were rinsed in poultry rinse bags with 400 ml of Butterfield's phosphate buffer in which Salmonella was enumerated from the diluents and Butterfield's phosphate. Chicxide significantly reduced Salmonella by 1.3 log CFU/ml with spray treatment and 2.3 log CFU/ml for all dip treatments. Following 30 min of attachment, the beef tips were placed into a spray cabinet with either Beefxide or sterilized water (control) and sprayed at 1 ft/2.5 s chain speed at 40 lb/in(2). The external surface of each beef tip was swabbed (100 cm(2)) to determine pathogen loads. Beefxide significantly reduced E. coli O157:H7 by 1.4 log CFU/100 cm(2) and Salmonella by 1.1 log CFU/100 cm(2) (P < 0.05) compared with the control samples.

Evaluation of nonpathogenic surrogate bacteria as process validation indicators for Salmonella enterica for selected antimicrobial treatments, cold storage, and fermentation in meat.

Prerigor lean and adipose beef carcass tissues were artificially inoculated individually with stationary-phase cultures of five nonpathogenic Escherichia coli cultures that had been previously identified as surrogates for E. coli O157:H7 or a mixture of five Salmonella strains in a fecal inoculum. Each tissue sample was processed with microbial interventions comparable with those used in the meat industry. The log reductions of the E. coli isolates were generally not statistically different from the salmonellae inoculum within a specific treatment. Inoculation experiments were also conducted with ground beef stored at either 4 or -20 degrees C. When compared with the Salmonella inoculum, at least three of the five E. coli strains survived in a manner that was not statistically different from the salmonellae. The E. coli strains and the Salmonella mixed culture were also inoculated into summer sausage batter, and the population enumerated both before and after fermentation. Four of the E. coli strains showed a lower population reduction (higher survival) than the Salmonella mixed culture. The five nonpathogenic E. coli strains may be used as individually or collectively for specific process validation indicators for Salmonella.