Biodistribution and Clearance of Stable Superparamagnetic Maghemite Iron Oxide Nanoparticles in Mice Following Intraperitoneal Administration.

Nanomedicine is an emerging field with great potential in disease theranostics. We generated sterically stabilized superparamagnetic iron oxide nanoparticles (s-SPIONs) with average core diameters of 10 and 25 nm and determined the in vivo biodistribution and clearance profiles. Healthy nude mice underwent an intraperitoneal injection of these s-SPIONs at a dose of 90 mg Fe/kg body weight. Tissue iron biodistribution was monitored by atomic absorption spectroscopy and Prussian blue staining. Histopathological examination was performed to assess tissue toxicity. The 10 nm s-SPIONs resulted in higher tissue-iron levels, whereas the 25 nm s-SPIONs peaked earlier and cleared faster. Increased iron levels were detected in all organs and body fluids tested except for the brain, with notable increases in the liver, spleen, and the omentum. The tissue-iron returned to control or near control levels within 7 days post-injection, except in the omentum, which had the largest and most variable accumulation of s-SPIONs. No obvious tissue changes were noted although an influx of macrophages was observed in several tissues suggesting their involvement in s-SPION sequestration and clearance. These results demonstrate that the s-SPIONs do not degrade or aggregate in vivo and intraperitoneal administration is well tolerated, with a broad and transient biodistribution. In an ovarian tumor model, s-SPIONs were shown to accumulate in the tumors, highlighting their potential use as a chemotherapy delivery agent.

The biology of meniscal pathology in osteoarthritis and its contribution to joint disease: beyond simple mechanics.

The meniscal cartilages in the knee function to improve congruity of the medial and lateral femoro-tibial joints and play critical roles in load distribution and joint stability. Meniscal tears of various configurations are one of the most common conditions of the knee and are associated with an increased risk of developing osteoarthritis (OA). While this risk has been largely attributed to loss of the biomechanical functions of the menisci, there is accumulating evidence suggesting that other aspects of meniscal biology may play a role in determining the long-term consequences of meniscal damage for joint health. In this narrative review, we examine the existing literature and present some new data implicating synthesis and secretion of enzymes and other pro-catabolic mediators by injured and degenerate menisci, contributing to the pathological change in other knee joint tissues in OA.

1H NMR spectroscopy of serum reveals unique metabolic fingerprints associated with subtypes of surgically induced osteoarthritis in sheep.

Osteoarthritis (OA) is a highly prevalent joint disease. Its slow progressive nature and the correlation between pathological changes and clinical symptoms mean that OA is often well advanced by the time of diagnosis. In the absence of any specific pharmacological treatments, there is a pressing need to develop robust biomarkers for OA. We have adopted a nuclear magnetic resonance (NMR)-based metabolomic strategy to identify molecular responses to surgically induced OA in an animal model. Sheep underwent one of three types of surgical procedure (sham (control), meniscal destabilization, MD or anterior cruciate ligament transaction, ACLT), and for every animal a serum sample was collected both pre- and postoperatively, thus, affording two types of "control" data for comparison. 1D 1H NMR spectra were acquired from each sample at 800 MHz and the digitized spectral data were analyzed using principal components analysis and partial least-squares regression discriminant analysis. Our approach, combined with the study design, allowed us to separate the metabolic responses to surgical intervention from those associated with OA. We were able to identify dimethyl sulfone (DMSO2) as being increased in MD after 4 weeks, while ACLT-induced OA exhibited increased 3-methylhistidine and decreased branched chain amino acids (BCAAs). The findings are discussed in the context of interpretation of metabolomic results in studies of human disease, and the selection of appropriate "control" data sets.

Acticoat versus biobrane: a retrospective review on the treatment of paediatric mid-dermal torso burns.

OBJECTIVES: Scalds involving toddlers commonly involve the torso and are frequently mid-dermal in depth. Initial management of a mid-dermal burn is conservative, progressing to grafting if healing has not been achieved in 10-14 days. Historically BiobraneTM (UDL Laboratories, Inc., Sugar Land, TX) is thought to have more favourable clinical outcomes compared to Acticoat TM (Smith and Nephew, St. Petersburg, Fl, USA). The Burns Unit at The Children's Hospital at Westmead (CHW) uses both dressings on a regular basis, providing the opportunity to compare the results of the dressings in a cohort of patients with mid-dermal torso burns. METHOD: A retrospective review was undertaken of all paediatric mid-dermal torso burns admitted to CHW between 2015 and 2017. The primary outcomes analysed were: time to complete healing and the need for grafting. Secondary outcomes included: operating theatre time, clinic visits, length of stay in hospital and positive wound swab colonisation. RESULTS: 78 children met the study criteria, 64 (82%) in the Acticoat group and 14 (18%) in the Biobrane group. 36 out of 78 children (56%) in the Acticoat group had their burns spontaneously healed without the need of skin graft surgery, compared with 10 out of 14 children (71%) in the Biobrane group. The days to complete healing were quicker in the Acticoat group (13 days) compared to the Biobrane group (17 days), although this was not statistically significant (P = 0.3). Overall patients managed with the Biobrane dressing required more operative sessions under general anaesthesia, a longer hospital stay, more clinic visits and a higher number of positive wound swab colonisation with heavy growth compared to the Acticoat group. CONCLUSION: This study suggests that the use of the Biobrane dressing does not significantly improve the clinical outcomes of mid-dermal torso burns in children compared to the Acticoat dressing. Acticoat reduced healing time, decreased the requirements for a general anaesthesia, reduced inpatient hospital stay and risk of infection.

Hyaluronan oligosaccharides stimulate matrix metalloproteinase and anabolic gene expression in vitro by intervertebral disc cells and annular repair in vivo.

The role of hyaluronan (HA) oligosaccharides in disc cell-mediated matrix metalloproteinase (MMP) and anabolic gene expression in vitro and annular repair in vivo were examined. Monolayer and alginate bead cultures of ovine intervertebral disc cells were stimulated with 10-12 mer hyaluronan oligosaccharides (HA-oligos). Annulus fibrosus (AF) monolayers were poorly responsive to the HA-oligos, proMMP-2 levels were marginally elevated and levels were MMP-9 unaffected. ProMMP-2 displayed a strong dose-dependent increase in the nucleus pulposus (NP) monolayers. In AF alginate bead cultures, proMMP-2 and active MMP-9 increased up to day 10, in NP cultures proMMP-2 was progressively converted to active MMP-2 over days 7-10 and active MMP-9 levels were elevated on day 10. A steady decline in MMP-2 and MMP-9 activity was evident over days 2-10 in the non-stimulated NP cultures. Disc cell viabilities were ≥92 ± 5% in all cultures indicating that the HA-oligo was not cytotoxic. Reverse-transcription polymerase chain reaction demonstrated an upregulation in MMP1, MMP113 and ADAMTS1 and the anabolic matrix repair genes ACAN, COL1A1 and COL2A1 in the NP by HA-oligos, whereas AF MMP13, ADAMTS1, ADAMTS4 and ADAMTS5, ACAN and COL2A1 were down-regulated; this differential regulation is expected to promote clearance of granulation/scar tissue from AF defects and matrix replenishment. The AF defect sites contained enlarged annular lamellae in vivo in response to the HA oligos, which is consistent with an active repair response. Masson trichrome and PicroSirius red histology and immunolocalization of type I collagen supported active remodelling in the outer lesion zone by the HA-oligo treatment but not the inner lesion. Copyright © 2016 John Wiley & Sons, Ltd.

Biglycan and fibromodulin fragmentation correlates with temporal and spatial annular remodelling in experimentally injured ovine intervertebral discs.

This study evaluated spatial and temporal extracellular matrix changes, induced by controlled surgical defects in the outer third of the annulus fibrosus (AF) of ovine intervertebral discs (IVDs). Thirty-two 4 year old sheep received a 4 mm deep x 10 mm wide standard annular surgical incision in the L1L2 and L3L4 IVDs (lesion group), 32 sheep were also subjected to the same surgical approach but the AF was not incised (sham-operated controls). Remodeling of the IVD matrix in the lesion and sham discs was assessed histochemically at 3, 6,12 and 26 month post operation (PO). Discs were also dissected into annular lesion site and contra-lateral AF and NP and equivalent zones in the sham sheep group, extracted with GuHCl, dialysed, freeze dried, digested with chondroitinase ABC/keratanase-I and aliquots examined for small leucine repeat proteoglycan (SLRP) core protein species by Western blotting using C-terminal antibodies to decorin, biglycan, lumican and fibromodulin and monoclonal antibody (Mab) 2B6 to unsaturated stub epitopes on chondroitin-4-sulphate generated by chondroitinase ABC. Masson Trichrome and Picrosirius red staining demonstrated re-organisation of the outermost collagenous lamellae in the incised discs 3-6 month PO. Toluidine blue staining also demonstrated a focal loss of anionic proteoglycan (PG) from the annular lesion 3-6 month PO with partial recovery of PG levels by 26 month. Specific fragments of biglycan and fibromodulin were associated with remodeling of the AF 12-26 month PO in the lesion IVDs but were absent from the NP of the lesion discs or all tissue zones in the sham animal group. Fragments of decorin were also observed in lesion zone extracts from 3 to 6 months but diminished after this. Isolation and characterization of the biglycan/fibromodulin fragments may identify them as prospective biomarkers of annular remodeling and characterization of the enzyme systems responsible for their generation may identify therapeutic target molecules.

Implementation of Epic Beaker Anatomic Pathology at an Academic Medical Center.

BACKGROUND: Beaker is a relatively new laboratory information system (LIS) offered by Epic Systems Corporation as part of its suite of health-care software and bundled with its electronic medical record, EpicCare. It is divided into two modules, Beaker anatomic pathology (Beaker AP) and Beaker Clinical Pathology. In this report, we describe our experience implementing Beaker AP version 2014 at an academic medical center with a go-live date of October 2015. METHODS: This report covers preimplementation preparations and challenges beginning in September 2014, issues discovered soon after go-live in October 2015, and some post go-live optimizations using data from meetings, debriefings, and the project closure document. RESULTS: We share specific issues that we encountered during implementation, including difficulties with the proposed frozen section workflow, developing a shared specimen source dictionary, and implementation of the standard Beaker workflow in large institution with trainees. We share specific strategies that we used to overcome these issues for a successful Beaker AP implementation. Several areas of the laboratory-required adaptation of the default Beaker build parameters to meet the needs of the workflow in a busy academic medical center. In a few areas, our laboratory was unable to use the Beaker functionality to support our workflow, and we have continued to use paper or have altered our workflow. In spite of several difficulties that required creative solutions before go-live, the implementation has been successful based on satisfaction surveys completed by pathologists and others who use the software. However, optimization of Beaker workflows has continued to be an ongoing process after go-live to the present time. CONCLUSIONS: The Beaker AP LIS can be successfully implemented at an academic medical center but requires significant forethought, creative adaptation, and continued shared management of the ongoing product by institutional and departmental information technology staff as well as laboratory managers to meet the needs of the laboratory.

Granulosa Cell-Specific Brca1 Loss Alone or Combined with Trp53 Haploinsufficiency and Transgenic FSH Expression Fails to Induce Ovarian Tumors.

BRCA1 mutations are associated with ovarian cancer. Previous studies reported that murine granulosa cell (GC) Brca1 loss caused ovarian-uterine tumors resembling serous cystadenomas, but the pathogenesis of these tumors may have been confounded by ectopic Brca1 expression and altered estrous cycling. We have used Tg.AMH.Cre conferring proven ovarian and GC-specific Cre activity to selectively target Brca1 disruption, denoted Brca1(GC-/-). Furthermore, ovary-specific Brca1(GC-/-) was combined with global Trp53 haploinsufficiency (Trp53(+/-)) and transgenic follicle-stimulating hormone (Tg.FSH) overexpression as a multi-hit strategy to investigate additional genetic and hormonal ovarian tumorigenesis mechanisms. However, 12-month-old Brca1(GC-/-) mice had no detectable ovarian or uterine tumors. Brca1(GC-/-) mice had significantly increased ovary weights, follicles exhibiting more pyknotic granulosa cells, and fewer corpora lutea with regular estrous cycling compared to controls. Isolated Brca1(GC-/-) mutation lengthened the estrous cycle and proestrus stage; however, ovarian cystadenomas were not observed, even when Brca1(GC-/-) was combined with Trp53(+/-) and overexpressed Tg.FSH. Our Brca1(GC-/-) models reveal that specific intra-follicular Brca1 loss alone, or combined with cancer-promoting genetic (Trp53 loss) and endocrine (high serum FSH) changes, was not sufficient to cause ovarian tumors. Our findings show that the ovary is remarkably resistant to oncogenesis, and support the emerging view of an extragonadal, multi-hit origin for ovarian tumorigenesis.

Fragmentation of decorin, biglycan, lumican and keratocan is elevated in degenerate human meniscus, knee and hip articular cartilages compared with age-matched macroscopically normal and control tissues.

INTRODUCTION: The small leucine-rich proteoglycans (SLRPs) modulate tissue organization, cellular proliferation, matrix adhesion, growth factor and cytokine responses, and sterically protect the surface of collagen type I and II fibrils from proteolysis. Catabolism of SLRPs has important consequences for the integrity of articular cartilage and meniscus by interfering with their tissue homeostatic functions. METHODS: SLRPs were dissociatively extracted from articular cartilage from total knee and hip replacements, menisci from total knee replacements, macroscopically normal and fibrillated knee articular cartilage from mature age-matched donors, and normal young articular cartilage. The tissue extracts were digested with chondroitinase ABC and keratanase-I before identification of SLRP core protein species by Western blotting using antibodies to the carboxyl-termini of the SLRPs. RESULTS: Multiple core-protein species were detected for all of the SLRPs (except fibromodulin) in the degenerate osteoarthritic articular cartilage and menisci. Fibromodulin had markedly less fragments detected with the carboxyl-terminal antibody compared with other SLRPs. There were fewer SLRP catabolites in osteoarthritic hip than in knee articular cartilage. Fragmentation of all SLRPs in normal age-matched, nonfibrillated knee articular cartilage was less than in fibrillated articular cartilage from the same knee joint or total knee replacement articular cartilage specimens of similar age. There was little fragmentation of SLRPs in normal control knee articular cartilage. Only decorin exhibited a consistent increase in fragmentation in menisci in association with osteoarthritis. There were no fragments of decorin, biglycan, lumican, or keratocan that were unique to any tissue. A single fibromodulin fragment was detected in osteoarthritic articular cartilage but not meniscus. All SLRPs showed a modest age-related increase in fragmentation in knee articular and meniscal cartilage but not in other tissues. CONCLUSION: Enhanced fragmentation of SLRPs is evident in degenerate articular cartilage and meniscus. Specific decorin and fibromodulin core protein fragments in degenerate meniscus and/or human articular cartilage may be of value as biomarkers of disease. Once the enzymes responsible for their generation have been identified, further research may identify them as therapeutic targets.