Sub-1.4 cm3 capsule for detecting labile inflammatory biomarkers in situ.

Transient molecules in the gastrointestinal tract such as nitric oxide and hydrogen sulfide are key signals and mediators of inflammation. Owing to their highly reactive nature and extremely short lifetime in the body, these molecules are difficult to detect. Here we develop a miniaturized device that integrates genetically engineered probiotic biosensors with a custom-designed photodetector and readout chip to track these molecules in the gastrointestinal tract. Leveraging the molecular specificity of living sensors1, we genetically encoded bacteria to respond to inflammation-associated molecules by producing luminescence. Low-power electronic readout circuits2 integrated into the device convert the light emitted by the encapsulated bacteria to a wireless signal. We demonstrate in vivo biosensor monitoring in the gastrointestinal tract of small and large animal models and the integration of all components into a sub-1.4 cm3 form factor that is compatible with ingestion and capable of supporting wireless communication. With this device, diseases such as inflammatory bowel disease could be diagnosed earlier than is currently possible, and disease progression could be more accurately tracked. The wireless detection of short-lived, disease-associated molecules with our device could also support timely communication between patients and caregivers, as well as remote personalized care.

Our surgical technique to treat premature growth plate closure 7 years after toe-to-thumb transfer. A case report.

In children, traumatic distal amputations of the thumb can be treated by partial first toe transfer. Growth is preserved by conserving a portion of the growth plate in the hallux distal phalanx. In the patient featured here, 7 years after such a distal thumb reconstruction, bone bridge resection was needed to restart growth and correct clinodactyly. When this patient was reviewed 4 years later, the thumb's longitudinal growth had been restored and continued.

Do teenagers return to normal physical activity levels after limb fractures? A longitudinal, accelerometry-based, activity monitoring study.

PURPOSE: This study aimed to elucidate whether levels of physical activity (PA) return to normal after bone healing or whether long-term behavioural changes in PA are to be expected in children and teenagers who have sustained limb fractures. METHODS: In all, 100 children and teenagers with a first episode of limb fracture and 100 sex- and age-matched healthy controls (CTRL) were recruited for a prospective study. PA in limb fracture patients was assessed at 18-month follow-up using accelerometer measurements, and values were compared with those of CTRL. Time spent in PA at different levels of intensity was determined for each participant and expressed in minutes and as a percentage of total validly measured time. RESULTS: Mean levels of PA at different levels of intensity by previously injured children and teenagers were similar than CTRL (42 sets of paired data). However, time spent in moderate-to-vigorous PA (MVPA) was lower than 60 minutes among limb-fracture patients at 18-month follow-up. CONCLUSION: The amount of skeletal loading in children and teenagers returns to normal values by 18 months after limb fracture. Even if time spent in MVPA is not significantly lower in children and teenagers with limb fractures, it no longer reached the international recommendations for school-aged children (MVPA > 60 minutes), which may be interpreted as a lifestyle modification or a behavioural change to avoid new trauma. LEVEL OF EVIDENCE: II.

A new technique for correction of leg length discrepancies in combination with complex axis deformities of the lower limb using a lengthening nail and a locking plate.

PURPOSE: Intramedullary lengthening nails are an accepted alternative to external fixators but are limited by anatomical preconditions. Therefore, to date the use of external fixators is sometimes inevitable. We report on a new technique for correction of combined limb length discrepancies and complex axis deformities using solely internal devices - a lengthening nail and a locking plate. METHODS: Between October 2008 and November 2011 five patients (two femora, three tibias) with a mean leg length discrepancy of 36 mm (25 to 50) and a complex angular deformity were treated with a fully implantable motorized lengthening nail (Fitbone) and a locking plate. All patients were evaluated with regards to the pre- and postoperative leg length as well as axis alignment, functional outcome, lengthening indices and complications. RESULTS: A successful leg length equalization was achieved in all cases and physiological joint orientation angles in all but one case. The mean distraction index was 1.2 mm/day, the maturation index 24 days/cm and the consolidation index 35 days/cm. The functional outcome was very encouraging in all cases with bilateral free range of movement. In total, two complications were observed, one nonunion and one loss of leg length after an early locking bolt removal in a peripheral hospital. CONCLUSION: The combination of a fully implantable motorized lengthening nail and a locking plate is a valuable alternative option for treating selected cases with limb length discrepancies in combination with a complex deformity of the lower leg. However, the reported technique puts high demands on the preoperative planning, operative technique as well as surgeon's skills. LEVEL OF EVIDENCE: IV (retrospective series).

The role of the periosteal sleeve in the reconstruction of bone defects using a non-vascularised fibula graft in the pediatric population.

BACKGROUND: Following resection of large benign bone tumors surgeons are confronted with bone defects severely affecting the stability of a limb. To restore the mechanical continuity of the bone different treatment methods using bone grafts have been described. In pediatric patients the thick periosteal sleeve is thought to contribute to bone formation. HYPOTHESIS: An intact periosteal sleeve is crucial in bone remodelling around a non-vascularised fibular graft used to bridge large bone defects. METHODS: We present a treatment technique applied in 6 cases comprising of subperiosteal tumor resection at the diaphyseal or metaphyseal level of long bones followed by defect bridging with a non-vascularised fibula graft inserted into the periosteal sleeve of the resection zone. Elastic intramedullary nails or plates were used for stabilisation. RESULTS: Due to the intact periosteum at the resection site bone integration occurred quickly and full remodelling was seen in all but one case. Tumor location in this case was at the metaphyseal level resulting in tumor resection at the growth plate. Although bone healing at the distal resection site was seen after a few weeks proximal consolidation was only partial. Full reconstitution of the fibula in the remaining periosteal sleeve was seen in 5 cases, partial reconstitution in 1 case. DISCUSSION: In the pediatric patient, the described technique is an effective and reliable treatment method for large benign bone tumors requiring resection. However, great diameter discrepancy of the donor and recipient site and a thin periosteum can be a limiting factor for its application. LEVEL OF EVIDENCE: Level IV clinical study.

Nestin-expressing cells in the gut give rise to enteric neurons and glial cells.

BACKGROUND: Neuronal stem cells (NSCs) are promising for neurointestinal disease therapy. Although NSCs have been isolated from intestinal musclularis, their presence in mucosa has not been well described. Mucosa-derived NSCs are accessible endoscopically and could be used autologously. Brain-derived Nestin-positive NSCs are important in endogenous repair and plasticity. The aim was to isolate and characterize mucosa-derived NSCs, determine their relationship to Nestin-expressing cells and to demonstrate their capacity to produce neuroglial networks in vitro and in vivo. METHODS: Neurospheres were generated from periventricular brain, colonic muscularis (Musc), and mucosa-submucosa (MSM) of mice expressing green fluorescent protein (GFP) controlled by the Nestin promoter (Nestin-GFP). Neuronal stem cells were also grown as adherent colonies from intestinal mucosal organoids. Their differentiation potential was assessed using immunohistochemistry using glial and neuronal markers. Brain and gut-derived neurospheres were transplanted into explants of chick embryonic aneural hindgut to determine their fate. KEY RESULTS: Musc- and MSM-derived neurospheres expressed Nestin and gave rise to cells of neuronal, glial, and mesenchymal lineage. Although Nestin expression in tissue was mostly limited to glia co-labelled with glial fibrillary acid protein (GFAP), neurosphere-derived neurons and glia both expressed Nestin in vitro, suggesting that Nestin+/GFAP+ glial cells may give rise to new neurons. Moreover, following transplantation into aneural colon, brain- and gut-derived NSCs were able to differentiate into neurons. CONCLUSIONS & INFERENCES: Nestin-expressing intestinal NSCs cells give rise to neurospheres, differentiate into neuronal, glial, and mesenchymal lineages in vitro, generate neurons in vivo and can be isolated from mucosa. Further studies are needed for exploring their potential for treating neuropathies.

Full-field optical coherence microscopy is a novel technique for imaging enteric ganglia in the gastrointestinal tract.

BACKGROUND: Noninvasive methods are needed to improve the diagnosis of enteric neuropathies. Full-field optical coherence microscopy (FFOCM) is a novel optical microscopy modality that can acquire 1 µm resolution images of tissue. The objective of this research was to demonstrate FFOCM imaging for the characterization of the enteric nervous system (ENS). METHODS: Normal mice and EdnrB(-/-) mice, a model of Hirschsprung's disease (HD), were imaged in three-dimensions ex vivo using FFOCM through the entire thickness and length of the gut. Quantitative analysis of myenteric ganglia was performed on FFOCM images obtained from whole-mount tissues and compared with immunohistochemistry imaged by confocal microscopy. KEY RESULTS: Full-field optical coherence microscopy enabled visualization of the full thickness gut wall from serosa to mucosa. Images of the myenteric plexus were successfully acquired from the stomach, duodenum, colon, and rectum. Quantification of ganglionic neuronal counts on FFOCM images revealed strong interobserver agreement and identical values to those obtained by immunofluorescence microscopy. In EdnrB(-/-) mice, FFOCM analysis revealed a significant decrease in ganglia density along the colorectum and a significantly lower density of ganglia in all colorectal segments compared with normal mice. CONCLUSIONS & INFERENCES: Full-field optical coherence microscopy enables optical microscopic imaging of the ENS within the bowel wall along the entire intestine. FFOCM is able to differentiate ganglionic from aganglionic colon in a mouse model of HD, and can provide quantitative assessment of ganglionic density. With further refinements that enable bowel wall imaging in vivo, this technology has the potential to revolutionize the characterization of the ENS and the diagnosis of enteric neuropathies.

The phosphorycholine moiety of the filarial nematode immunomodulator ES-62 is responsible for its anti-inflammatory action in arthritis.

OBJECTIVE: In countries where parasitic infections are endemic, autoimmune disease is relatively rare, leading to the hypothesis that parasite-derived immunomodulators may protect against its development. Consistent with this, we have previously demonstrated that ES-62, a 62 kDa phosphorylcholine (PC)-containing glycoprotein that is secreted by filarial nematodes, can exert anti-inflammatory action in the murine collagen-induced arthritis (CIA) model and human rheumatoid arthritis-derived synovial tissue cultures. As a first step to developing ES-62-based drugs, the aim of this study was to determine whether the PC-moiety of ES-62 was responsible for its anti-inflammatory actions. METHODS: We compared the anti-inflammatory activity of a PC-free form of recombinant ES-62 (rES-62) and a synthetic PC-ovalbumin conjugate (OVA-PC) with that of native ES-62 in the CIA model and synovial tissues from patients with rheumatoid arthritis. RESULTS: The anti-inflammatory actions of ES-62 in CIA appear to be dependent on the PC moiety as indicated by the reduction in severity of disease and also suppression of collagen-specific T helper 1 cytokine production observed when testing OVA-PC, but not rES-62. Interestingly, the anti-inflammatory activity of PC did not correlate with a reduction in anti-collagen IgG2a levels. Also, the ES-62-mediated suppression of interferon-gamma from human patient tissues could be mimicked by OVA-PC but not rES-62 or ovalbumin. CONCLUSIONS: In countries where filariasis is endemic the reduced detection of inflammatory diseases, such as rheumatoid arthritis may be because of the anti-inflammatory action of the PC moieties of ES-62. PC may thus provide the starting point for the development of novel, safe immunomodulatory therapies.

Cellular and complement-dependent cytotoxicity of Ep-CAM-specific monoclonal antibody MT201 against breast cancer cell lines.

MT201 is a fully human monoclonal IgG1 antibody with moderate affinity for epithelial cell adhesion molecule (Ep-CAM) being clinically developed for the treatment of carcinomas. Like many other clinically validated IgG1 monoclonal antibodies, MT201 primarily acts by antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Here, we analysed ADCC and CDC induced by MT201 and, as reference, trastuzumab against a panel of nine human breast cancer cell lines expressing distinct surface levels of Ep-CAM and human epithelial growth factor receptor type 2 antigen. Maximal cell lysis by ADCC by MT201 and trastuzumab in the presence of peripheral mononuclear cells did not significantly differ when averaged over the nine cell lines, but showed marked differences with respect to individual cell lines. The extent of cell lysis at intermediate surface target density was highly variable, suggesting a dominant influence of other susceptibility factors. Only one breast cancer cell line was eliminated via CDC, but only by MT201. Resistance to CDC appeared to correlate with high expression levels of complement resistance factors. Our present data as well as recent data on the prevalence and prognostic relevance of Ep-CAM expression in metastatic breast cancer suggest that Ep-CAM-specific monoclonal IgG1 antibodies may have a significant therapeutic potential in the treatment of breast cancer.

Determinants of viral clearance and persistence during acute hepatitis C virus infection.

The virological and immunological features of hepatitis C virus (HCV) infection were studied weekly for 6 months after accidental needlestick exposure in five health care workers, four of whom developed acute hepatitis that progressed to chronicity while one subject cleared the virus. In all subjects, viremia was first detectable within 1-2 weeks of inoculation, 1 month or more before the appearance of virus-specific T cells. The subject who cleared the virus experienced a prolonged episode of acute hepatitis that coincided with a CD38+ IFN-gamma- CD8+ T cell response to HCV and a small reduction in viremia. Subsequently, a strong CD4+ T cell response emerged and the CD8+ T cells became CD38- and started producing IFN-gamma in response to HCV, coinciding with a rapid 100,000-fold decrease in viremia that occurred without a corresponding surge of disease activity. Chronic infection developed in two subjects who failed to produce a significant T cell response and in two other subjects who initially mounted strong CD4+ T cell responses that ultimately waned. In all subjects, viremia was higher at the peak of acute hepatitis than it was when the disease began, and the disease improved during the viremia. These results provide the first insight into the host-virus relationship in humans during the incubation phase of acute HCV infection, and they provide the only insight to date into the virological and immunological characteristics of clinically asymptomatic acute HCV infection, the commonest manifestation of this disease. In addition, the results suggest that the vigor and quality of the antiviral T cell response determines the outcome of acute HCV infection, that the ability of HCV to outpace the T cell response may contribute to its tendency to persist; that the onset of hepatitis coincides with the onset of the CD8+ T cell response, that disease pathogenesis and viral clearance are mediated by different CD8+ T cell populations that control HCV by both cytolytic and noncytolytic mechanisms, and that there are different pathways to viral persistence in asymptomatic and symptomatic acute HCV infection.

Regional differences in the distribution of catalase in the epithelium of the ocular lens.

Oxidative stress is thought to play a major role in cataract formation. The present experiments are aimed at gaining a better understanding of the systems that protect the lens from damage by reactive oxygen species. The aqueous humor normally contains hydrogen peroxide (H2O2), a compound capable of generating reactive oxygen species. The systems protecting the ocular lens from oxidative damage are primarily confined to the epithelium, a single layer of cells on the anterior side of the organ directly beneath the lens capsule. When cultured rabbit lenses were challenged with a single dose of 0.2 mM H2O2, cells in the peripheral region of the epithelium survived; those in the central region died. Here we investigate the histochemical and immunoperoxidase distributions of catalase, an enzyme which detoxifies H2O2, in cells from the peripheral and central regions of the epithelium on flat mount preparations of the epithelium. In a flat mount, the entire population of lens epithelial cells can be viewed on one preparation. The reaction product for catalase activity and its immunoperoxidase localization were more intense in peripheral epithelial cells than in cells throughout the central epithelium. Treatment of cultured lens epithelial cells or rabbit lenses with 3-aminotriazole or potassium cyanide, inhibitors of catalase, reduced or abolished the histochemical reaction product. Ultrastructural cytochemistry confirmed the presence of catalase in microperoxisomes of the epithelial cells from whole lenses. The decreased level of catalase throughout the central epithelium may account for the increased susceptibility of these cells to H2O2-induced cell death.

Prevalence of adoptees among special education populations.

A statewide survey of public and private schools was conducted to determine the prevalence of adoptees among children classified for educational purposes as neurologically impaired (NI), perceptually impaired (PI), or emotionally disturbed (ED). Results indicated that adopted children were overrepresented in these special education populations, accounting for 6.7% of NI students, 5.4% of PI students, and 7.2% of ED students. Implications of the findings for educational and clinical intervention are discussed.