Functional and histologic effects of a free nonvascularised muscle graft implanted into a reinnervated muscle after prolonged denervation

Striated muscle atrophy and degeneration, which increase with the delay of denervation, represent two of the main causes for poor recovery following delayed nerve repair. The present study, using a rat model, tests the hypothesis that an adjunction of small, free, nonvascularised muscle grafts of contralateral healthy triceps into a chronically denervated triceps improves muscle regeneration and recovery following sciatic nerve repair delayed for 3 months. Our experiments seem to show a relative increase in mechanical properties in animals in which free muscle graft into the triceps was performed 3 weeks following nerve repair. The improvement of the regenerative process of muscles which have suffered a long period of denervation should be considered as an additional therapeutic procedure in the case of late nerve repair.     

Unmyelinated nerve fiber analysis of the human abducent nerve

Unmyelinated nerve fibers of the abducent nerve have occasionally been observed with electron microscopes, but, to our knowledge, they have never been evaluated from the morphometric point of view. We analyzed the aging process of the unmyelinated nerve fibers in the human abducent nerve with the help of a new staining method suitable for morphometric research on the nervous system. We studied numbers and transverse areas of unmyelinated fibers of the abducent nerve in 10 cadavers. Our findings were that (1) these fibers were distributed diffusely, (2) their number decreased with age, and (3) the mean transverse area did not change with age. Most of the unmyelinated axons were thinner than the myelinated axons. These results may be important for analysis of clinical signs in relation to aging and ophthalmologic functions.     

Fusimotor-free spindles in reinnervated muscles of neonatal rats treated with nerve growth factor

Crushing the nerve to the medial gastrocnemius muscle in newborn rats and administering nerve growth factor afterwards results in a reinnervated muscle containing supernumerary muscle spindles. The structure and innervation of 88 spindles in the reinnervated muscles were reconstructed from serial thick and thin transverse sections at 30-35 days after the nerve crush, and compared to those of five control spindles. The spindles consisted of one to four small-diameter encapsulated fibers with features of nuclear chain intrafusal fibers, or infrequently a nuclear bag intrafusal fiber. Some of the spindles were located within a capsule that also contained an extrafusal fiber. Each spindle was innervated by an afferent with features of the primary afferent. The density of secondary afferents was lower in reinnervated muscles than in controls. Endplates were observed on extrafusal fibers in the experimental muscles, attesting to restoration of skeletomotor (alpha) innervation after the nerve crush. However, 78% of the experimental spindles were entirely devoid of efferent innervation. The remainder received either one or two fusimotor (gamma) axons or a skeletofusimotor (beta) axon, compared to the six to eight motor axons that innervated control spindles. The presence of supernumerary spindles composed of fibers that resemble normal intrafusal fibers in the absence of motor innervation suggests that afferents alone can induce the formation and subsequent differentiation of intrafusal fibers in nerve-crushed muscles of neonatal rats. In addition, the paucity of gamma innervation in nerve-crushed muscles suggests that immature gamma neurons are more susceptible than spindle afferents or alpha efferents to cell death after axotomy at birth.     

Intraoperative measurement of choline acetyltransferase activity to evaluate the functional status of donor nerve during reinnervated free muscle transfer: a preliminary report

Intraoperative choline acetyltransferase activity measurements were used to evaluate the functional status of donor nerves during reinnervated free muscle transfer. This technique was applied to 3 cases. One was a lower-type brachial plexus injury in combination with radial nerve injury; the other 2 were Volkmann's ischemic contractures. Gracilis muscle was transferred for reconstruction of wrist extension in all cases. The donor nerves included 1 anterior interosseous nerve and 2 posterior interosseous nerves. A fascicle with choline acetyltransferase activity above 2,000 cpm was considered to be reliable as a motor fascicle. Reinnervation of transferred muscle was confirmed by electromyographic examination within the first 3.3 months (range, 2.5 to 4 months) after surgery and all muscles obtained useful recovery. This technique can directly and quantitatively verify the functional quality of the donor motor fascicle when the quality of the donor nerve is in doubt.     

Myelinated nerve fibre analysis of the human small splanchnic nerve

The aim of the present study is to analyse nerve fibres of the human small splanchnic nerve in relation to the ageing process. The analysis was conducted with the use of a new staining method that makes it possible to discriminate various structures of the nervous tissue. An image-analysing digitiser, a microscope with a drawing tube and a personal computer for storing data and performing statistical analyses were employed in this study. We examined 30 human small splanchnic nerves which were taken from cadaver specimens (20 males and 10 females) aged from 44 to 96 years. Our report may provide a first information concerning the ageing process of the human small splanchnic nerve. The results indicate that a decrease in transverse area and perimeter of myelinated axons in one of the important changes occurring in the human small splanchnic nerve with the ageing process.     

Quantitative study of muscle fibre atrophy and restitution after nerve grafts

Our comparative experimental studies on rabbits using clinical, electromyographical, and quantitative histological examinations of long autologous and homologous nerve grafts in 35 rabbits, paying special attention to quantitative histological changes in the gastrocnemius muscles, allow the statement that, compared to the short holografts, the long homografts showed worse results. Moreover, it became obvious that the regeneration rate of autografts was not influenced by increasing the lengths of the grafts. The answer to the question of how far even longer grafts may influence the quality of regeneration will need further investigations.     

Expression of different isoforms of nitric oxide synthase in experimentally denervated and reinnervated skeletal muscle

Denervated muscle fibers express enhanced levels of stress and apoptosis-associated proteins and undergo apoptosis. In experimentally denervated and reinnervated rat facial muscle, we now evaluate changes in the expression patterns of different isoforms of nitric oxide synthase (NOS)-generating nitric oxide (NO), which mediates oxidative stress and apoptosis. Physiological expression of NOS corresponds to a constant sarcolemmal staining pattern for neuronal NOS (nNOS) and a patchy sarcolemmal and weak sarcoplasmic labeling for the endothelial NOS-isoform, with no expression for inducible NOS (iNOS). Denervated muscle displayed distinct downregulation of nNOS with preserved expression of dystrophin. Also, denervated and immediately reinnervated muscle fibers showed decreased expression of nNOS. However, muscle fibers reinnervated for 10 weeks revealed a restored physiological expression of nNOS. There were no changes in the expression of endothelial and inducible NOS. As NO is known to induce growth arrest and collapse of neuronal growth cones, downregulation of NOS may contribute to promotion of axonal regeneration by aiding formation of new endplates. NO is upregulated in reinnervated muscle fibers and thus prevents polyneural hyperinnervation by extrajunctional synapses. Furthermore, downregulation of NOS during denervation is compatible with the finding that low levels of NO contribute to apoptosis instead of necrosis in disease states of oxidative stress.     

Morphometric nerve fiber analysis and aging process of the human abducent nerve

BACKGROUND: Coronary artery disease (CAD) constitutes a considerable financial burden on society in Finland; it is the cause of death of approximately 7,500 men and 6,500 women annually in a population of 5 million. OBJECTIVES: The purpose of this study was to assess the changes in the quality of life (QOL) of patients with CAD treated by medication, percutaneous transluminal coronary angioplasty (PTCA), and coronary artery bypass surgery (CABS) during 1 year. METHOD: The study population consisted of 280 patients with CAD. One hundred patients had been referred to CABS and another 100 to PTCA, whereas 80 patients were on drug therapy. The patients assessed their health status and QOL in terms of functional capacity and aspects of distress using self-completed questionnaires with the Nottingham Health Profile (NHP) instrument before the operation and 6and 12 months afterwards. RESULTS: The QOL of the patients who had undergone CABS and PTCA was significantly better on the dimensions of energy, pain, and mobility 1 year after the intervention. In the medication group, the only improvement took place on the dimension of social isolation, whereas both energy and mobility deteriorated. CONCLUSIONS: The results on QOL obtained in this study support the notion that patients continue to have many problems even after medical treatment with a good outcome. The problems occur in different areas compared with the pretreatment situation as on the dimensions of social isolation and emotional reaction. The rehabilitation of CAD patients is therefore important because the new problems are manifested differently from those seen before the illness or the treatment. The patient's QOL and personal preference for a treatment modality should be important criteria in the choice of treatment.     

Tubular repair of the median nerve in the human forearm. Preliminary findings

Transected median nerves in the forearm of two male patients, 12 and 21 years of age, were treated with a chamber technique leaving a 3 to 5 mm gap between the nerve ends. The nerve ends were enclosed in a silicone tube of such a dimension that would not cause compression of the nerve. Post-operative examination including sensory evaluation and assessment of muscle contraction force was carried out after 3 years. In both cases there was excellent motor recovery of the thenar muscles. Outgrowth of sensory fibres was remarkably fast, resulting ultimately in functional sensibility allowing almost normal hand function. 2PD was < or = 6 mm (12-year-old patient) and 8 to 10 mm (21-year-old patient) respectively. In one case the silicone tube was re-explored because of minor local discomfort 2 years after the repair. The former gap was bridged by a smooth continuous nerve-like structure of the same diameter as the adjacent nerve trunk and with no signs of neuroma formation or compression of the nerve.     

Full functional recovery after surgical repair of transected abducens nerve: case report

OBJECTIVE AND IMPORTANCE: Results of surgical repair of the injured abducens nerve are rarely reported in the literature. A full functional recovery of a completely resected abducens root may be exceptional. We describe a patient who obtained normal ocular alignment and binocular vision after surgical reconstruction of a transected abducens nerve. CLINICAL PRESENTATION: A 56-year-old woman with a petroclival meningioma was presented. She underwent total removal of the tumor through a combined supra/infratentorial transpetrosal approach. The abducens nerve was tightly attenuated by the tumor and thickened dura. During dissection, the nerve was completely transected just behind the entrance to Dorello's canal. INTERVENTION: The abducens nerve was the single root type and inevitably required surgical repair. To obtain a sufficient length of the distal stump for trimming, part of the petrosphenoidal ligament was cut and the superior border of the petrous bone was exposed. The proximal stump of the nerve was also trimmed to obtain healthy tissue, and reconstruction was performed with five 10-0 nylon sutures. Five months later, esodeviation began to improve. Nine months after the surgery, the patient did not complain of diplopia and an objective assessment reported normal ocular alignment and estimated binocular function as "excellent" according to Biglan's system. Overcorrection of abduction did not occur. CONCLUSION: The result in our patient confirms the possibility of full functional recovery after surgical repair of a totally transected abducens nerve.     

Gait analysis of adult paraplegic rats after spinal cord repair

This study presents a novel detailed method of analysis of rat gait and uses this method to demonstrate recovery of forward locomotion patterns in adult rats made paraplegic by surgical spinal cord transection and subjected to a novel strategy for spinal cord repair. Six normal rats were compared to five animals in which the cord was transected at T8-T9, and a 5-mm segment of the spinal cord removed, and to seven animals in which, following spinal cord transection and removal of a spinal cord segment, multiple intercostal peripheral nerve bridges were implanted, rerouting pathways from white to gray matter in both directions. The implanted area was filled with fibrin glue containing acidic fibroblast growth factor. Details of the repair strategy have been published (H. Cheng, Y. Cao, and L. Olson, 1996, Science 273: 510-513). Gait analysis was carried out 3 and 4 months after surgery and once in the normal animals. Animals were allowed to walk across a runway with a transparent floor. Each test consisted of five trials, and each trial was videorecorded from underneath. Using frame-by-frame playback, individual footprints were then recorded regarding location and order of limb use, as well as step quality (degree of weight bearing, etc.). These data allowed measuring runway transit time, five different measures of step numbers, all possible temporal patterns of limb use, stride length, and base of support. Transected controls remained paralyzed in the hindlimbs with only occasional reflex hindlimb movements without weight bearing. Animals subjected to the full repair procedure were significantly faster than the controls, used their hindlimbs for 25-30% of the movements, and regained several of the specific limb recruitment patterns used by normal rats. Taken together, the gait analysis data demonstrate remarkable recovery of coordinated gait in the repaired animals, which was significantly better than controls for all relevant parameters, while at the same time clearly inferior to normal rats for most of the examined parameters. We conclude that normal rats use a multitude of interchangeable step sequence patterns, and that our spinal cord repair strategy leads to recovery of some of these patterns following complete spinal cord transection. These data suggest functionally relevant neuronal communication across the lesion.     

Promoter analysis of the human mismatch repair gene hMSH2

On the basis of both the interleukin-2-receptor (IL-2R) alpha-chain expression on 16-day-old fetal rat thymocytes and the occurrence of interleukin-2 (IL-2) mRNA-containing cells early during rat thymus ontogeny, we have investigated the possible role of IL-2/IL-2R complex in rat T-cell maturation. For this purpose, we analyzed the effects of the addition of either recombinant rat IL-2 or anti-CD25 (OX-39)-blocking monoclonal antibodies to fetal thymus organ cultures (FTOC), established from 16-day-old rat embryos. IL-2 stimulated the growth of thymocytes and, as a result, induced T-cell differentiation, whereas OX-39 mAb blocked the maturation of thymic-cell progenitors. Accordingly, these results support the involvement of IL-2/IL-2R complex in rat T-cell development.     

Influence of collagen and laminin gels concentration on nerve regeneration after resection and tube repair

In order to assess the usefulness of collagen and laminin gels prefilling nerve chambers to enhance nerve regeneration, we compared reinnervation of target organs after sciatic nerve resection leaving gaps of 4 or 6 mm followed by repair with silicone tubes in different groups of mice. Tubes were prefilled with saline solution, collagen gels, or laminin-containing gels at different concentrations. Functional reinnervation was assessed by noninvasive methods to quantitate recovery of sweating, nociceptive, sensory, and motor functions in the hindpaw repeatedly during 4-5 months postoperation. The increase in gap length between nerve stumps delayed the beginning and reduced the degree of functional recovery achieved. Reinnervation started earlier and achieved slightly higher levels with collagen gel diluted at 1.28 mg/ml than with more concentrated (1.92 and 2.56 mg/ml) collagen gels and with saline-prefilled tubes bridging a 4-mm gap. Recovery was also better with diluted (4 mg/ml) than with concentrated (12 mg/ml) laminin-containing gel, although lower than with collagen gels and saline. By prefilling silicone tubes bridging a 6-mm gap, a length considered limiting for regeneration in the mouse sciatic nerve, with diluted collagen or laminin gels, both matrices allowed for higher levels of recovery and for successful regeneration in a higher proportion of mice than saline solution. The laminin gel performed slightly better than the collagen gel.     

Myosin heavy chain isoforms of human muscle after short-term spaceflight

The influence of microgravity on the myosin phenotype of skeletal muscle fibers in the vastus lateralis of eight crew members was studied before and after 5-day (n = 3) and 11-day (n = 5) spaceflights (space shuttle flights: STS-32, -33 and -34). Single-fiber electrophoresis analyses showed that the proportion of fibers expressing only slow (type I) myosin heavy chain (MHC) in the vastus lateralis was significantly lower after than before 11 days of spaceflight. Although the family of type II MHC isoforms was elevated post- compared with preflight, the distribution among the isoforms of type II MHC was not statistically different. Based on monoclonal and polyclonal antibodies specific for three adult MHC isoforms and single-fiber electrophoresis, approximately 3% of the fibers analyzed coexpressed all three adult MHC isoforms. The results from immunohistochemical staining with two different sets of antibodies indicate a reduction in the percentage of fibers expressing type I MHC as a result of spaceflight. The mean difference, however, was significant only when the fibers were categorized simply as type I or II. These changes appeared to be highly individualized among the astronauts. These results suggest that a rapid change in MHC isoform expression can occur in some muscle fibers after a relatively brief exposure to spaceflight.     

Functional analysis of human replication protein A in nucleotide excision repair

Human replication protein A (RPA) is a three-subunit protein complex (70-, 34-, and 11-kDa subunits) involved in DNA replication, repair, and recombination. Both the 70- (p70) and 34-kDa (p34) subunits interact with Xeroderma pigmentosum group A complementing protein (XPA), a key protein involved in nucleotide excision repair. Our deletion analysis indicated that no particular domain(s) of RPA p70 was essential for its interaction with XPA, whereas 33 amino acids from the C terminus of p34 (p34Delta33C) were necessary for the XPA interaction. Furthermore, mutant RPA lacking the p34 C terminus failed to interact with XPA, suggesting that p34, not p70, is primarily responsible for the interaction of RPA with XPA. RPA stimulated the interaction of XPA with UV-damaged DNA through an RPA-XPA complex on damaged DNA sites because (i) the RPA mutant lacking the C terminus of p34 failed to stimulate an XPA-DNA interaction, and (ii) the ssDNA binding domain of RPA (amino acids 296-458) was necessary for the stimulation of the XPA-DNA interaction. Two separate domains of p70, a single-stranded DNA binding domain and a zinc-finger domain, were necessary for RPA function in nucleotide excision repair. The mutant RPA (RPA:p34Delta33C), which lacks its stimulatory effect on the XPA-DNA interaction, also poorly supported nucleotide excision repair, suggesting that the XPA-RPA interaction on damaged DNA is necessary for DNA repair activity.     

An experimental study on the recovery of the lingual nerve after injury with or without repair

BACKGROUND: The advantage of radionuclide angiographic techniques used to measure right ventricular ejection fraction (RVEF) is geometry independence, but the weakness is right atrial (RA) overlap. To minimize the effect of RA counts on right ventricular time activity curve (TAC), two regions of interest (ROI), one drawn for the end-diastolic image and one for the end-systolic image, are used for the calculation of RVEF from equilibrium gated blood pool scans (GBPS) and from gated first-pass studies with an Anger camera. A multicrystal camera offers both temporal separation of the bolus to the right side of the heart and good count statistics; therefore first-pass studies performed on a multicrystal camera theoretically should yield the most accurate measurements of RVEF, but few studies have been performed to validate RVEF against a reliable gold standard. METHODS AND RESULTS: To develop and validate an accurate method to measure RVEF from multicrystal first-pass data, 25 patients underwent sequential cine-MRI, first-pass radionuclide angiography, and gated equilibrium imaging. Five additional healthy volunteers underwent cine-MRI alone. Right and left ventricular volumes were measured from serial short axis cine-MRI views according to Simpson's rule. Three methods were used to calculate RVEF from first-pass data: a single ROI method, a dual ROI method, and a method in which a single ROI is applied to RA subtracted first-pass dynamic data. Five additional healthy volunteers underwent cine-MRI alone. When right ventricular stroke volume was plotted versus left ventricular stroke volume for the 5 volunteers and the 15 patients without valvular regurgitation, the regression line was not significantly different from the line of identity, supporting the accuracy of cine-MRI to measure RVEF. The RVEF by cine-MRI ranged from 34% to 59%; first-pass RVEF with a single ROI from 26% to 48%; first-pass RVEF with two ROIs from 31% to 59%; first-pass RVEF with RA subtracted single ROI from 29% to 60%; and RVEF from GBPS with multiple ROIs from 28% to 55%. The regressions for all three of the first-pass methods versus cine-MRI were significant (p < 0.01) as was the regression for the equilibrium GBPS versus cine-MRI but the correlation was weaker. The regressions for the 2-ROI method and for the RA subtracted single ROI method were not significantly different from the line of identity, whereas the regressions for both the single ROI method and for equilibrium GBPS were significantly different from the line of identity (p < 0.01). CONCLUSIONS: Cine-MRI can be used to validate radionuclide algorithms. Of the four radionuclide methods for measuring RVEF that were assessed, the first-pass 2-ROI method and the first-pass RA subtracted single ROI are the most accurate, the first-pass single ROI method underestimates RVEF, and the RVEF values measured from GBPS are less accurate.