Expression of Semaphorins, Neuropilins, VEGF, and Tenascins in Rat and Human Primary Sensory Neurons after a Dorsal Root Injury.

Dorsal root injury is a situation not expected to be followed by a strong regenerative growth, or growth of the injured axon into the central nervous system of the spinal cord, if the central axon of the dorsal root is injured but of strong regeneration if subjected to injury to the peripherally projecting axons. The clinical consequence of axonal injury is loss of sensation and may also lead to neuropathic pain. In this study, we have used in situ hybridization to examine the distribution of mRNAs for the neural guidance molecules semaphorin 3A (SEMA3A), semaphorin 3F (SEMA3F), and semaphorin 4F (SEMA4F), their receptors neuropilin 1 (NP1) and neuropilin 2 (NP2) but also for the neuropilin ligand vascular endothelial growth factor (VEGF) and Tenascin J1, an extracellular matrix molecule involved in axonal guidance, in rat dorsal root ganglia (DRG) after a unilateral dorsal rhizotomy (DRT) or sciatic nerve transcetion (SNT). The studied survival times were 1-365 days. The different forms of mRNAs were unevenly distributed between the different size classes of sensory nerve cells. The results show that mRNA for SEMA3A was diminished after trauma to the sensory nerve roots in rats. The SEMA3A receptor NP1, and SEMA3F receptor NP2, was significantly upregulated in the DRG neurons after DRT and SNT. SEMA4F was upregulated after a SNT. The expression of mRNA for VEGF in DRG neurons after DRT showed a significant upregulation that was high even a year after the injuries. These data suggest a role for the semaphorins, neuropilins, VEGF, and J1 in the reactions after dorsal root lesions.

Epidemiology and changed surgical treatment methods for fractures of the distal radius: a registry analysis of 42,583 patients in Stockholm County, Sweden, 2004­2010.

BACKGROUND AND PURPOSE: The incidence of fractures of the distal radius may have changed over the last decade, and operative treatment has been commoner during that time. We investigated the incidence of fractures of the distal radius and changing trends in surgical treatment during the period 2004-2010. PATIENTS AND METHODS: Registry data on 42,583 patients with a fracture of the distal radius from 2004 to 2010 were evaluated regarding diagnosis, age, sex, and surgical treatment. RESULTS: The crude incidence rate was 31 per 10(4) person-years with a bimodal distribution. After the age of 45 years, the incidence rate in women increased rapidly and leveled off first at a very high age. The incidence rate in postmenopausal women was lower than previously reported. In men, the incidence was low and it increased slowly until the age of 80 years, when it amounted to 31 per 10(4) person-years. The number of surgical procedures increased by more than 40% despite the fact that there was reduced incidence during the study period. In patients ≥ 18 years of age, the proportion of fractures treated with plating increased from 16% to 70% while the use of external fixation decreased by about the same amount. INTERPRETATION: The incidence rate of distal radius fractures in postmenopausal women appears to have decreased over the last few decades. There has been a shift in surgical treatment from external fixation to open reduction and plating.

Secretagogin is expressed in sensory CGRP neurons and in spinal cord of mouse and complements other calcium-binding proteins, with a note on rat and human.

BACKGROUND: Secretagogin (Scgn), a member of the EF-hand calcium-binding protein (CaBP) superfamily, has recently been found in subsets of developing and adult neurons. Here, we have analyzed the expression of Scgn in dorsal root ganglia (DRGs) and trigeminal ganglia (TGs), and in spinal cord of mouse at the mRNA and protein levels, and in comparison to the well-known CaBPs, calbindin D-28k, parvalbumin and calretinin. Rat DRGs, TGs and spinal cord, as well as human DRGs and spinal cord were used to reveal phylogenetic variations. RESULTS: We found Scgn mRNA expressed in mouse and human DRGs and in mouse ventral spinal cord. Our immunohistochemical data showed a complementary distribution of Scgn and the three CaBPs in mouse DRG neurons and spinal cord. Scgn was expressed in ~7% of all mouse DRG neuron profiles, mainly small ones and almost exclusively co-localized with calcitonin gene-related peptide (CGRP). This co-localization was also seen in human, but not in rat DRGs. Scgn could be detected in the mouse sciatic nerve and accumulated proximal to its constriction. In mouse spinal cord, Scgn-positive neuronal cell bodies and fibers were found in gray matter, especially in the dorsal horn, with particularly high concentrations of fibers in the superficial laminae, as well as in cell bodies in inner lamina II and in some other laminae. A dense Scgn-positive fiber network and some small cell bodies were also found in the superficial dorsal horn of humans. In the ventral horn, a small number of neurons were Scgn-positive in mouse but not rat, confirming mRNA distribution. Both in mouse and rat, a subset of TG neurons contained Scgn. Dorsal rhizotomy strongly reduced Scgn fiber staining in the dorsal horn. Peripheral axotomy did not clearly affect Scgn expression in DRGs, dorsal horn or ventral horn neurons in mouse. CONCLUSIONS: Scgn is a CaBP expressed in a subpopulation of nociceptive DRG neurons and their processes in the dorsal horn of mouse, human and rat, the former two co-expressing CGRP, as well as in dorsal horn neurons in all three species. Functional implications of these findings include the cellular refinement of sensory information, in particular during the processing of pain.

Selectivity in the reinnervation of the lateral gastrocnemius muscle after nerve repair with ethyl cyanoacrylate in the rat.

There is a need for complementary surgical techniques that enable rapid and reliable primary repair of transected nerves. Previous studies after peripheral nerve transection and repair with synthetic adhesives have demonstrated regeneration to an extent comparable to that of conventional techniques. The aim of this study was to compare two different repair techniques on the selectivity of muscle reinnervation after repair and completed regeneration. We used the cholera toxin B technique of retrograde axonal tracing to evaluate the morphology, the number, and the three-dimensional location of α-motoneurons innervating the lateral gastrocnemius muscle and compared the results after repair with either ethyl cyanoacrylate (ECA) or epineural sutures of the transected parent sciatic nerve. In addition, we recorded the wet weight of the muscle. Six months after transection and repair of the sciatic nerve, the redistribution of the motoneuron pool was markedly disorganized, the motoneurons had apparently increased in number, and they were scattered throughout a larger volume of the spinal cord gray matter with a decrease in the synaptic coverage compared to controls. A reduction in muscle weight was observed as well. No difference in morphometric variables or muscle weight between the two repair methods could be detected. We conclude that the selectivity of motor reinnervation following sciatic nerve transection and subsequent repair with ECA is comparable to that following conventional micro suturing.

Phospholipase C{beta}3 in mouse and human dorsal root ganglia and spinal cord is a possible target for treatment of neuropathic pain.

Treatment of neuropathic pain is a major clinical problem. This study shows expression of phospholipase ss3 (PLCss3) in mouse and human DRG neurons, mainly in small ones and mostly with a nonpeptidergic phenotype. After spared nerve injury, the pain threshold was strongly reduced, and systemic treatment of such animals with the unselective PLC inhibitor U73122 caused a rapid and long-lasting (48-h) increase in pain threshold. Thus, inhibition of PLC may provide a way to treat neuropathic pain.

Integrin-laminin interactions controlling neurite outgrowth from adult DRG neurons in vitro.

A prerequisite for axon regeneration is the interaction between the growth cone and the extracellular matrix (ECM). Laminins are prominent constituents of ECM throughout the body, known to support axon growth in vitro and in vivo. The regenerative capacity of adult neurons is greatly diminished compared to embryonic or early postnatal neurons. Since most lesions in the nervous system occur in the adult, we have examined neurite outgrowth from adult mouse DRG neurons on four laminin isoforms (laminin-1/LM-111, laminin-2/LM-211, laminin-8/LM-411 and laminin-10/LM-511) in vitro. The growth on laminin-1 and -10 was trophic factor-independent and superior to the one on laminin-2 and -8, where growth was very poor in the absence of neurotrophins. Among other ECM proteins, laminins were by far the most active molecules. Using function-blocking antibodies to laminin-binding integrins, we identified non-overlapping functions of integrins alpha3beta1, alpha7beta1 and alpha6beta1 on different laminin isoforms, in that alpha3beta1 and alpha7beta1 integrins appeared to be specific receptors for both laminin-1 and-2, whereas integrin alpha6beta1 was a receptor for laminin-8 and-10. Lastly, by use of immunohistochemistry, expression of subunits of laminin-1, -2, -8 and -10 in sensory organs in the human epidermis could be demonstrated, supporting an important role for these laminins in relation to primary sensory axons.

Impeded interaction between Schwann cells and axons in the absence of laminin alpha4.

The Schwann cell basal lamina (BL) is required for normal myelination. Loss or mutations of BL constituents, such as laminin-2 (alpha2beta1gamma1), lead to severe neuropathic diseases affecting peripheral nerves. The function of the second known laminin present in Schwann cell BL, laminin-8 (alpha4beta1gamma1), is so far unknown. Here we show that absence of the laminin alpha4 chain, which distinguishes laminin-8 from laminin-2, leads to a disturbance in radial sorting, impaired myelination, and signs of ataxia and proprioceptive disturbances, whereas the axonal regenerative capacity is not influenced. In vitro studies show poor axon growth of spinal motoneurons on laminin-8, whereas it is extensive on laminin-2. Schwann cells, however, extend longer processes on laminin-8 than on laminin-2, and, in contrast to the interaction with laminin-2, solely use the integrin receptor alpha6beta1 in their interaction with laminin-8. Thus, laminin-2 and laminin-8 have different critical functions in peripheral nerves, mediated by different integrin receptors.

Integrin messenger RNAs in the red nucleus after axotomy and neurotrophic administration.

Integrins are cell surface receptors known to be important for regeneration in the peripheral nervous system. We have investigated the expression of integrin messenger RNAs in red nucleus neurons of adult rats after axotomy and administration of neurotrophic factors. Using radioactive in situ hybridization, messenger RNA for integrin subunits beta1, alpha3, alpha7 and alphaV could be detected. No change of any alpha subunit could be detected after axotomy. In contrast, a small upregulation of beta1 was detected after lesion. Administration of neurotrophin-3 induced a robust further increase in beta1 messenger RNA levels, whereas brain-derived neurotrophic factor did not. By analogy to the peripheral nervous system, we propose that integrins may be important for a regenerative response in central nervous system neurons.

Dorsal root ganglion neurons up-regulate the expression of laminin-associated integrins after peripheral but not central axotomy.

The favorable prognosis of regeneration in the peripheral nervous system after axonal lesions is generally regarded as dependent on the Schwann cell basal lamina. Laminins, a heterotrimeric group of basal lamina molecules, have been suggested to be among the factors playing this supportive role. For neurons to utilize laminin as a substrate for growth, an expression of laminin binding receptors, integrins, is necessary. In this study, we have examined the expression of laminin binding integrin subunits in dorsal root ganglion (DRG) neurons after transection to either their peripherally projecting axons, as in the sciatic nerve, followed by regeneration, or the centrally projecting axons in dorsal roots, followed by no or weak regenerative activity. In uninjured DRG, immunohistochemical staining revealed a few neurons expressing integrin subunit alpha6, whereas integrin subunits alpha7 and foremost beta1 were expressed in a majority of neurons. After an injury to the sciatic nerve, mRNAs encoding all three integrins were up-regulated in DRG neurons. By anterograde tracing, immunoreactivity for all studied integrins was also found in association with growing axons after a sciatic nerve crush lesion in vivo. In contrast, mRNA levels remained constant in DRG neurons after a dorsal root injury. Together with previous findings, this suggests that integrin subunits alpha6, alpha7, and beta1 have an important role in the regenerative response following nerve injury and that the lack of regenerative capacity following dorsal root injury could in part be explained by the absence of response in integrin regulation.

Induction of HIF1alpha but not HIF2alpha in motoneurons after ventral funiculus axotomy-implication in neuronal survival strategies.

Spinal cord injury is frequently associated with local tissue hypoxia. As neuronal cells are susceptible to damage caused by low oxygen levels, hypoxia-induced activation of tissue-protective factors could represent an endogenous mechanism for neuron survival following injury. We studied in vivo, in a rat model of intraspinal axotomy of motoneurons, the cell- and time-dependent regulation of the hypoxia-inducible transcription factors (HIFs), HIF1alpha and HIF2alpha, as well as one of their target genes, vascular endothelial growth factor (VEGF). VEGF is a potent hypoxia-regulated angiogenic growth factor with recently discovered neuroprotective and neurotrophic activities. While neither HIF1alpha, HIF2alpha, nor VEGF mRNA were detected in noninjured motoneurons, we found a strong induction of HIF1alpha, but not HIF2alpha mRNA in axotomized motoneurons. HIF1alpha expression peaked at about 7 days after injury. Moreover, we found increased VEGF mRNA and protein expression around and within the scar but also within motoneurons, peaking around 3 days after axotomy. In addition, increased survival of cultured motoneurons after treatment with VEGF could also be shown. We conclude that axotomized motoneurons in this model respond to injury by specific induction of HIF1alpha and VEGF expression that may provide an endogenous mechanism with the potential to promote motoneuron survival after injury.

Laminin chains in rat and human peripheral nerve: distribution and regulation during development and after axonal injury.

During nerve growth, axons are dependent upon contact with matrix components, such as laminins, for elongation, guidance, and trophic support. Semiquantitative in situ hybridization histochemistry and immunohistochemistry (IHC) were used to identify laminin chains in normal peripheral nerves, during postnatal development, after sciatic nerve transection (SNT), and after sciatic nerve crush (SNC). Laminin alpha2, alpha4, beta1, beta2, and gamma1 chain mRNAs were all expressed at high levels in newborn rat sciatic nerves with declining levels during later developmental stages. At the adult stage, no laminin chain mRNA was detectable. Of interest, the mRNA levels for alpha4 chain declined faster than those for alpha2. After SNT, laminin alpha2, alpha4, beta1, and gamma1 mRNA levels were up-regulated at the site of the injury, with the most profound reaction in the proximal nerve stump. Laminin alpha2 and alpha4 chains differed in that the mRNA levels of alpha4 were up-regulated earlier and declined quicker, whereas alpha2 had a later onset, with high levels remaining even after 6 weeks. After SNC, there was an initial up-regulation of the same laminin chain mRNAs as after SNT in the nerve, however, less intense, and at 6 weeks after SNC, all laminin mRNA levels studied had returned to normal. IHC of adult human normal and transected peripheral nerves stained positive for laminin alpha2, alpha4, beta1, and gamma1 chains in close relation to neurofilament labeled axons. Laminin alpha3, alpha4, alpha5, beta1, beta2, and gamma1 chains were found in blood vessel-like structures and alpha3, alpha4, alpha5, beta2, and gamma1 in the perineurium. These results and a previously published description of integrin regulation in spinal motoneurons suggest that both laminin-2 (alpha2beta1gamma1) and laminin-8 (alpha4beta1gamma1) are important for the postnatal nerve development and axonal regeneration after injury and that laminin-8 may have important functions especially early postnatally and early after adult nerve lesion.