Upper Extremity Wood Saw Injuries: Experience at a Single Level 1 Trauma Center.

BACKGROUND: Saw injuries are common, with more 75 000 occurring each year in the United States alone. While these injuries occur frequently, management strategies are not universally agreed upon, and data regarding outcomes and complications are lacking. We propose to provide a comprehensive picture of upper extremity saw injury patterns, management strategies, complications, and outcomes. METHODS: Patients presenting to a single level 1 trauma center between 2012 and2019 with upper extremity laceration, crush, or amputation were identified. In all, 10 721 patients were reviewed, and those without wood saw injuries were excluded. Patient demographic information, injury details, management strategy, and outcomes were collected. RESULTS: In all, 283 upper extremity wood saw injuries were analyzed. Injuries most commonly affected the fingers (92.2%), and the frequencies of simple lacerations and complicated injuries were nearly identical. The table saw was the most commonly implicated saw (48%) with more than half of the injuries being complicated, the most common being bone injury. Most patients were treated nonsurgically (81.3%), with the majority undergoing wound care in the emergency department followed by home antibiotics (68.2%). Subsequent complications were exceedingly rare (4.2%), with wound infection occurring in 5 patients. Amputations occurred in 19.4% of patients, leading to permanent functional impairment. CONCLUSIONS: Wood saw injuries are common, generating functional and financial burden. While injuries range in severity, management can typically be conducted within the emergency department with local wound care and outpatient oral antibiotics. Injury complications and long-term issues are rare. Ongoing efforts to promote saw safety are required to minimize the burden of these injuries.

Fyn-binding protein ADAP supports actin organization in podocytes.

The renal podocyte is central to the filtration function of the kidney that is dependent on maintaining both highly organized, branched cell structures forming foot processes, and a unique cell-cell junction, the slit diaphragm. Our recent studies investigating the developmental formation of the slit diaphragm identified a novel claudin family tetraspannin, TM4SF10, which is a binding partner for ADAP (also known as Fyn binding protein Fyb). To investigate the role of ADAP in podocyte function in relation to Fyn and TM4SF10, we examined ADAP knockout (KO) mice and podocytes. ADAP KO mice developed glomerular pathology that began as hyalinosis and progressed to glomerulosclerosis, with aged male animals developing low levels of albuminuria. Podocyte cell lines established from the KO mice had slower attachment kinetics compared to wild-type cells, although this did not affect the total number of attached cells nor the ability to form focal contacts. After attachment, the ADAP KO cells did not attain typical podocyte morphology, lacking the elaborate cell protrusions typical of wild-type podocytes, with the actin cytoskeleton forming circumferential stress fibers. The absence of ADAP did not alter Fyn levels nor were there differences between KO and wild-type podocytes in the reduction of Fyn activating phosphorylation events with puromycin aminonucleoside treatment. In the setting of endogenous TM4SF10 overexpression, the absence of ADAP altered the formation of cell-cell contacts containing TM4SF10. These studies suggest ADAP does not alter Fyn activity in podocytes, but appears to mediate downstream effects of Fyn controlled by TM4SF10 involving actin cytoskeleton organization.

Vertebrate Claudin/PMP22/EMP22/MP20 family protein TMEM47 regulates epithelial cell junction maturation and morphogenesis.

BACKGROUND: TMEM47 is the vertebrate orthologue of C. elegans VAB-9, a tetraspan adherens junction protein in the PMP22/EMP/Claudin family of proteins. VAB-9 regulates cell morphology and adhesion in C. elegans and TMEM47 is expressed during kidney development and regulates the activity of Fyn. The conserved functions of VAB-9 and TMEM47 are not well understood. RESULTS: expression of TMEM47 in C. elegans functionally rescues vab-9 mutations. Unlike Claudins, expression of TMEM47 in L fibroblasts does not generate tight junction strands; instead, membrane localization requires E-cadherin expression. Temporally, TMEM47 localizes at cell junctions first with E-cadherin before ZO-1 colocalization and in polarized epithelia, TMEM47 colocalizes with adherens junction proteins. By immunoprecipitation, TMEM47 associates with classical adherens junction proteins, but also with tight junction proteins Par6B and aPKCλ. Over-expression of TMEM47 in MDCK cells decreases apical surface area, increases activated myosin light chain at cell-cell contacts, disrupts cell polarity and morphology, delays cell junction reassembly following calcium switch, and selectively interferes with tight junction assembly. Reduced TMEM47 expression results in opposite phenotypes. CONCLUSIONS: TMEM47 regulates the localization of a subset of tight junction proteins, associated actomyosin structures, cell morphology, and participates in developmental transitions from adherens to tight junctions. Developmental Dynamics 245:653-666, 2016. © 2016 Wiley Periodicals, Inc.

The minus-end actin capping protein, UNC-94/tropomodulin, regulates development of the Caenorhabditis elegans intestine.

BACKGROUND: Tropomodulins are actin-capping proteins that regulate the stability of the slow-growing, minus-ends of actin filaments. The C. elegans tropomodulin homolog, UNC-94, has sequence and functional similarity to vertebrate tropomodulins. We investigated the role of UNC-94 in C. elegans intestinal morphogenesis. RESULTS: In the embryonic C. elegans intestine, UNC-94 localizes to the terminal web, an actin- and intermediate filament-rich structure that underlies the apical membrane. Loss of UNC-94 function results in areas of flattened intestinal lumen. In worms homozygous for the strong loss-of-function allele, unc-94(tm724), the terminal web is thinner and the amount of F-actin is reduced, pointing to a role for UNC-94 in regulating the structure of the terminal web. The non-muscle myosin, NMY-1, also localizes to the terminal web, and we present evidence that increasing actomyosin contractility by depleting the myosin phosphatase regulatory subunit, mel-11, can rescue the flattened lumen phenotype of unc-94 mutants. CONCLUSIONS: The data support a model in which minus-end actin capping by UNC-94 promotes proper F-actin structure and contraction in the terminal web, yielding proper shape of the intestinal lumen. This establishes a new role for a tropomodulin in regulating lumen shape during tubulogenesis.

The secretory pathway calcium ATPase PMR-1/SPCA1 has essential roles in cell migration during Caenorhabditis elegans embryonic development.

Maintaining levels of calcium in the cytosol is important for many cellular events, including cell migration, where localized regions of high calcium are required to regulate cytoskeletal dynamics, contractility, and adhesion. Studies show inositol-trisphosphate receptors (IP3R) and ryanodine receptors (RyR), which release calcium into the cytosol, are important regulators of cell migration. Similarly, proteins that return calcium to secretory stores are likely to be important for cell migration. The secretory protein calcium ATPase (SPCA) is a Golgi-localized protein that transports calcium from the cytosol into secretory stores. SPCA has established roles in protein processing, metal homeostasis, and inositol-trisphosphate signaling. Defects in the human SPCA1/ATP2C1 gene cause Hailey-Hailey disease (MIM# 169600), a genodermatosis characterized by cutaneous blisters and fissures as well as keratinocyte cell adhesion defects. We have determined that PMR-1, the Caenorhabditis elegans ortholog of SPCA1, plays an essential role in embryogenesis. Pmr-1 strains isolated from genetic screens show terminal phenotypes, such as ventral and anterior enclosure failures, body morphogenesis defects, and an unattached pharynx, which are caused by earlier defects during gastrulation. In Pmr-1 embryos, migration rates are significantly reduced for cells moving along the embryo surface, such as ventral neuroblasts, C-derived, and anterior-most blastomeres. Gene interaction experiments show changing the activity of itr-1/IP3R and unc-68/RyR modulates levels of embryonic lethality in Pmr-1 strains, indicating pmr-1 acts with these calcium channels to regulate cell migration. This analysis reveals novel genes involved in C. elegans cell migration, as well as a new role in cell migration for the highly conserved SPCA gene family.

Claudins reign: The claudin/EMP/PMP22/γ channel protein family in C. elegans.

The claudin family of integral membrane proteins was identified as the major protein component of the tight junctions in all vertebrates. Since their identification, claudins, and their associated pfam00822 superfamily of proteins have been implicated in a wide variety of cellular processes. Claudin homologs have been identified in invertebrates as well, including Drosophila and C. elegans. Recent studies demonstrate that the C. elegans claudins, clc-1-clc- 5, and similar proteins in the greater PMP22/EMP/claudin/voltage-gated calcium channel γ subunit family, including nsy-4, and vab-9, while highly divergent at a sequence level from each other and from the vertebrate claudins, in many cases play roles similar to those traditionally assigned to their vertebrate homologs. These include regulating cell adhesion and passage of small molecules through the paracellular space, channel activity, protein aggregation, sensitivity to pore-forming toxins, intercellular signaling, cell fate specification and dynamic changes in cell morphology. Study of claudin superfamily proteins in C. elegans should continue to provide clues as to how claudin family protein function has been adapted to perform diverse functions at specialized cell-cell contacts in metazoans.

TM4SF10 and ADAP interaction in podocytes: role in Fyn activity and nephrin phosphorylation.

TM4SF10 [transmembrane tetra(4)-span family 10] is a claudin-like cell junction protein that is transiently expressed during podocyte development where its expression is downregulated in differentiating podocytes coincident with the appearance of nephrin at the slit diaphragm. In a yeast two-hybrid screen, we identified adhesion and degranulation-promoting adaptor protein (ADAP), a well-known Fyn substrate and Fyn binding partner, as a TM4SF10 interacting protein in mouse kidney. Using coimmunoprecipitation and immunohistochemistry experiments in cultured human podocytes, we show that TM4SF10 colocalizes with Fyn and ADAP but does not form a stable complex with Fyn. Cytoskeletal changes and phosphorylation events mediated by Fyn activity were reversed by TM4SF10 overexpression, including a decrease in the activating tyrosine phosphorylation of Fyn (Y(421)), suggesting TM4SF10 may have a regulatory role in suppressing Fyn activity. In addition, TM4SF10 was reexpressed following podocyte injury by puromycin aminonucleoside treatment, and its expression enhanced the abundance of high-molecular-weight forms of nephrin indicating it may participate in a mechanism controlling nephrin's appearance at the plasma membrane. Therefore, these studies have identified ADAP as another Fyn adapter protein expressed in podocytes, and that TM4SF10, possibly through ADAP, may regulate Fyn activity. Since TM4SF10 expression is temporally regulated during kidney development, these studies may help define a mechanism by which the slit diaphragm matures as a highly specialized cell junction during podocyte differentiation.

Claudin family proteins in Caenorhabditis elegans.

In the last decade, the claudin family of integral membrane proteins has been identified as the major protein component of the tight junctions in all vertebrates. The claudin superfamily proteins also function to regulate channel activity, intercellular signaling, and cell morphology. Subsequently, claudin homologues have been identified in invertebrates, including Drosophila and Caenorhabditis elegans. Recent studies demonstrate that the C. elegans claudins, clc-1 to clc-5, and similar proteins in the greater PMP22/EMP/claudin/calcium channel γ subunit family, including nsy-1-nsy-4 and vab-9, while highly divergent at a sequence level from each other and from the vertebrate claudins, in some cases play roles similar to those traditionally assigned to their vertebrate homologues. These include regulating cell adhesion and passage of small molecules through the paracellular space. The claudin superfamily proteins also function to regulate channel activity, intercellular signaling, and cell morphology. Study of claudin superfamily proteins in C. elegans should continue to provide clues as to how core claudin protein function can be modified to serve various specific roles at regions of cell-cell contact in metazoans.

Expression of TM4SF10, a Claudin/EMP/PMP22 family cell junction protein, during mouse kidney development and podocyte differentiation.

Cell junctions in the nephron are highly specialized to perform specific and distinct filtration and reabsorption functions. The mature kidney forms complex cell junctions including slit diaphragms that prevent the passage of serum proteins into the filtrate, and tubule cell junctions that regulate specific paracellular ion reuptake. We have investigated the expression of TM4SF10 (Trans-Membrane tetra(4)-Span Family 10) in mouse kidneys. TM4SF10 is the vertebrate orthologue of Caenorhabditis elegans VAB-9, a tetraspan adherens junction protein in the PMP22/EMP/Claudin family of proteins. We found that TM4SF10 localizes at the basal-most region of podocyte precursors before the capillary loop stage, at some tubule precursors, and at the ureteric bud junction with S-shaped bodies. Overall expression of TM4SF10 peaked at postnatal day 4 and was virtually absent in adult kidneys. The very limited expression of TM4SF10 protein that persisted into adulthood was restricted to a few tubule segments but remained localized to the basal region of lateral membranes. In undifferentiated cultured podocytes, TM4SF10 localized to the perinuclear region and translocated to the cell membrane after Cadherin appearance at cell-cell contacts. TM4SF10 colocalized with ZO1 and p120ctn in undifferentiated confluent podocytes and also colocalized with the tips of actin filaments at cell contacts. Upon differentiation of cultured podocytes, TM4SF10 protein disappeared from cell contacts and expression ceased. These results suggest that TM4SF10 functions during differentiation of podocytes and may participate in the maturation of cell junctions from simple adherens junctions to elaborate slit diaphragms. TM4SF10 may define a new class of Claudin-like proteins that function during junctional development.

The inositol 1,4,5-trisphosphate receptor regulates epidermal cell migration in Caenorhabditis elegans.

Polarized migration and spreading of epithelial sheets is important during many processes in vivo, including embryogenesis and wound healing. However, the signaling pathways that regulate epithelial migrations are poorly understood. To identify molecular components that regulate the spreading of epithelial sheets, we performed a screen for mutations that perturb epidermal cell migration during embryogenesis in Caenorhabditis elegans. We identified one mutant (jc5) as a weak mutation in itr-1, which encodes the single inositol 1,4,5-trisphosphate receptor (ITR) in C. elegans. During the migration of the embryonic epidermis, jc5 embryos display defects including misdirected migration or premature cessation of migration. Cells that halt their migration have disorganized F-actin and display reduced filopodial protrusive activity at their leading edge. Furthermore, some filopodia formed by epidermal cells in itr-1(jc5) embryos exhibit abnormally long lifetimes. Pharmacological studies with the inositol 1,4,5-trisphosphate antagonist xestospongin C phenocopy these defects, confirming that ITR function is important for proper epidermal migration. Our results provide the first molecular evidence that movements of embryonic epithelial cell sheets can be controlled by ITRs and suggest that such regulation may be a widespread mechanism for coordinating epithelial cell movements during embryogenesis.

Nucleic acid-based techniques for post-transcriptional regulation of molecular targets.

PURPOSE OF REVIEW: Messenger RNA, transfer RNA and ribosomal RNA were defined long ago as essential components for transmission of genetic code from DNA. However, there are many other, less commonly recognized RNAs, such as ribozymes and small interfering RNAs, which are distinguished by their ability to inhibit RNA function. This review describes the basic molecular concepts and potential therapeutic applications of RNA inhibition by a variety of molecules, including ribozymes, antisense oligonucleotides, aptamers and small interfering RNAs. RECENT FINDINGS: A tremendous amount of data has recently emerged about double-stranded small interfering RNAs, which bind and degrade corresponding messenger RNAs by a process called RNA interference. Though native small interfering RNAs have been shown to be biologically relevant in animals and plants, synthetic types have rapidly become powerful tools for post-transcriptional inhibition of specific gene products to determine functional consequences in simple organisms and in-vitro model systems. More established means of RNA inhibition, such as with ribozyme and antisense strategies, continue to be viable options for in-vitro experiments, and form the basis for many ongoing clinical trials. SUMMARY: Ribozymes, antisense oligonucleotides, aptamers and small interfering RNAs are potentially useful reagents for in-vitro investigation and for treatment of kidney and hypertension diseases.

The cell junction protein VAB-9 regulates adhesion and epidermal morphology in C. elegans.

Epithelial cell junctions are essential for cell polarity, adhesion and morphogenesis. We have analysed VAB-9, a cell junction protein in Caenorhabditis elegans. VAB-9 is a predicted four-pass integral membrane protein that has greatest similarity to BCMP1 (brain cell membrane protein 1, a member of the PMP22/EMP/Claudin family of cell junction proteins) and localizes to the adherens junction domain of C. elegans apical junctions. Here, we show that VAB-9 requires HMR-1/cadherin for localization to the cell membrane, and both HMP-1/alpha-catenin and HMP-2/beta-catenin for maintaining its distribution at the cell junction. In vab-9 mutants, morphological defects correlate with disorganization of F-actin at the adherens junction; however, localization of the cadherin-catenin complex and epithelial polarity is normal. These results suggest that VAB-9 regulates interactions between the cytoskeleton and the adherens junction downstream of or parallel to alpha-catenin and/or beta-catenin. Mutations in vab-9 enhance adhesion defects through functional loss of the cell junction genes apical junction molecule 1 (ajm-1) and discs large 1 (dlg-1), suggesting that VAB-9 is involved in cell adhesion. Thus, VAB-9 represents the first characterized tetraspan adherens junction protein in C. elegans and defines a new family of such proteins in higher eukaryotes.