Increased Axon Initial Segment Length Results in Increased Na+ Currents in Spinal Motoneurones at Symptom Onset in the G127X SOD1 Mouse Model of Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease preferentially affecting motoneurones. Transgenic mouse models have been used to investigate the role of abnormal motoneurone excitability in this disease. Whilst an increased excitability has repeatedly been demonstrated in vitro in neonatal and embryonic preparations from SOD1 mouse models, the results from the only studies to record in vivo from spinal motoneurones in adult SOD1 models have produced conflicting findings. Deficits in repetitive firing have been reported in G93A SOD1(high copy number) mice but not in presymptomatic G127X SOD1 mice despite shorter motoneurone axon initial segments (AISs) in these mice. These discrepancies may be due to the earlier disease onset and prolonged disease progression in G93A SOD1 mice with recordings potentially performed at a later sub-clinical stage of the disease in this mouse. To test this, and to explore how the evolution of excitability changes with symptom onset we performed in vivo intracellular recording and AIS labelling in G127X SOD1 mice immediately after symptom onset. No reductions in repetitive firing were observed showing that this is not a common feature across all ALS models. Immunohistochemistry for the Na+ channel Nav1.6 showed that motoneurone AISs increase in length in G127X SOD1 mice at symptom onset. Consistent with this, the rate of rise of AIS components of antidromic action potentials were significantly faster confirming that this increase in length represents an increase in AIS Na+ channels occurring at symptom onset in this model.

Intramuscular Botulinum toxin A injections induce central changes to axon initial segments and cholinergic boutons on spinal motoneurones in rats.

Intramuscular injections of botulinum toxin block pre-synaptic cholinergic release at neuromuscular junctions producing a temporary paralysis of affected motor units. There is increasing evidence, however, that the effects are not restricted to the periphery and can alter the central excitability of the motoneurones at the spinal level. This includes increases in input resistance, decreases in rheobase currents for action potentials and prolongations of the post-spike after-hyperpolarization. The aim of our experiments was to investigate possible anatomical explanations for these changes. Unilateral injections of Botulinum toxin A mixed with a tracer were made into the gastrocnemius muscle of adult rats and contralateral tracer only injections provided controls. Immunohistochemistry for Ankyrin G and the vesicular acetylcholine transporter labelled axon initial segments and cholinergic C-boutons on traced motoneurones at 2 weeks post-injection. Soma size was not affected by the toxin; however, axon initial segments were 5.1% longer and 13.6% further from the soma which could explain reductions in rheobase. Finally, there was a reduction in surface area (18.6%) and volume (12.8%) but not frequency of C-boutons on treated motoneurones potentially explaining prolongations of the after-hyperpolarization. Botulinum Toxin A therefore affects central anatomical structures controlling or modulating motoneurone excitability explaining previously observed excitability changes.

Describing temporal variation in reticuloruminal pH using continuous monitoring data.

Reticuloruminal pH has been linked to subclinical disease in dairy cattle, leading to considerable interest in identifying pH observations below a given threshold. The relatively recent availability of continuously monitored data from pH boluses gives new opportunities for characterizing the normal patterns of pH over time and distinguishing these from abnormal patterns using more sensitive and specific methods than simple thresholds. We fitted a series of statistical models to continuously monitored data from 93 animals on 13 farms to characterize normal variation within and between animals. We used a subset of the data to relate deviations from the normal pattern to the productivity of 24 dairy cows from a single herd. Our findings show substantial variation in pH characteristics between animals, although animals within the same farm tended to show more consistent patterns. There was strong evidence for a predictable diurnal variation in all animals, and up to 70% of the observed variation in pH could be explained using a simple statistical model. For the 24 animals with available production information, there was also a strong association between productivity (as measured by both milk yield and dry matter intake) and deviations from the expected diurnal pattern of pH 2 d before the productivity observation. In contrast, there was no association between productivity and the occurrence of observations below a threshold pH. We conclude that statistical models can be used to account for a substantial proportion of the observed variability in pH and that future work with continuously monitored pH data should focus on deviations from a predictable pattern rather than the frequency of observations below an arbitrary pH threshold.

Immune evasion by acquisition of complement inhibitors: the mould Aspergillus binds both factor H and C4b binding protein.

Pathogenic fungi represent a major threat particularly to immunocompromised hosts, leading to severe, and often lethal, systemic opportunistic infections. Although the impaired immune status of the host is clearly the most important factor leading to disease, virulence factors of the fungus also play a role. Factor H (FH) and its splice product FHL-1 represent the major fluid phase inhibitors of the alternative pathway of complement, whereas C4b-binding protein (C4bp) is the main fluid phase inhibitor of the classical and lectin pathways. Both proteins can bind to the surface of various human pathogens conveying resistance to complement destruction and thus contribute to their pathogenic potential. We have recently shown that Candida albicans evades complement by binding both Factor H and C4bp. Here we show that moulds such as Aspergillus spp. bind Factor H, the splicing variant FHL-1 and also C4bp. Immunofluorescence and flow cytometry studies show that the binding of Factor H and C4bp to Aspergillus spp. appears to be even stronger than to Candida spp. and that different, albeit possibly nearby, binding moieties mediate this surface attachment.

Abnormal hepatic vein Doppler waveform in patients without liver disease.

In patients with liver cirrhosis Doppler ultrasound often detects absence of the retrograde (hepatopetal) flow phase in the hepatic vein, suggestive of an increased stiffness of the liver parenchyma around the vein. This is rarely or never reported in healthy control persons. We examined the frequency of absent retrograde flow in a consecutive series of 139 patients referred for abdominal ultrasound. We used state-of-the-art ultrasound scanners, and placed the Doppler gate so that in non-forced end-expiration it would sample the right hepatic vein 4-6 cm from the vena cava. There was no association between the hepatic vein flow pattern and age, sex or body mass index. 43 of 139 studied patients showed absent retrograde flow. Review of the case records revealed liver disease in 26 patients and no sign of liver disease in 17 patients. We suggest that absent retrograde flow in the hepatic veins may be seen not only in patients with overt liver disease but also in apparently liver-healthy patients.

Muscle reflexes during gait elicited by electrical stimulation of the posterior cruciate ligament in humans.

We investigated the influence of electrical stimulation of the posterior cruciate ligament (PCL) on the motoneuron pool of the thigh and calf muscle during gait. The study group comprised eight young men without any history of injury to the knee joints. Multistranded teflon-insulated stainless steel wires were inserted into the PCL guided by sonography and in four subjects also into the fat pad of the knee. The PCL was electrically stimulated during gait on a treadmill at heel strike and 100 ms after heel strike. Electromyographic signals were recorded with bipolar surface electrodes placed over the vastus medialis, rectus femoris, vastus lateralis, biceps femoris caput longum, and semitendinosus muscles. The stimuli consisted of four pulses delivered at 200 Hz; the stimulus amplitude was two to three times the sensory threshold. The electrical stimulation of the PCL inhibited the ongoing muscle activity in both the quadriceps and the hamstrings. The latency of the inhibition ranged between 78 and 148 ms in the quadriceps, between 88 and 110 ms in the hamstrings and between 189 and 258 ms in m. gastrocnemius. Stimulation of the fat pad of the knee did not influence the thigh and calf muscle motoneuron pool as evidenced by electromyography. The response elicited from the stimulation of the PCL was not limited to a specific muscle group but depended on ongoing muscle contraction, which suggests that the mechanoreceptors in the PCL are involved in the control of all muscles acting on the knee joint during gait.

Inhibition of dynamic thigh muscle contraction by electrical stimulation of the posterior cruciate ligament in humans.

We investigated the influence of electrical stimulation of the posterior cruciate ligament (PCL) on the motoneuron pool of the thigh muscle during voluntary static and dynamic muscle contraction. The study group comprised nine young men with no history of injury to the knee joints. Multistranded Teflon-insulated stainless-steel wires were inserted into the PCL guided by ultrasound. In three subjects wires were also inserted into the fat pad of the knee. The PCL was electrically stimulated during static, concentric, or eccentric muscle contraction with a constant load of 20% of the maximal voluntary contraction of either the quadriceps or the hamstrings. Electromyographic signals were recorded with bipolar surface electrodes placed over the vastus medialis, rectus femoris, vastus lateralis, biceps femoris caput longum, and semitendinosus muscles. The stimuli consisted of four pulses delivered at 200 HZ; the stimulus amplitude was two to three times the sensory threshold. The electrical stimulation of the PCL inhibited the ongoing muscle activity in both the quadriceps and hamstrings with latencies of 114-150 ms and 99-130 ms, respectively. Stimulation of the fat pad of the knee did not influence the muscle activity. The study suggests that the mechanoreceptors in the PCL are involved in controlling muscle activity during both static and active muscle contractions. The relative long latency of the reflex makes it unlikely that it can serve as a directly protective reflex for the cruciate ligaments.

Sonographic guided insertion of electrodes into the cruciate ligaments of the knee.

The present cadaver study examined the probability of accurate ultrasound guided insertion of electrodes into the anterior (ACL) and posterior (PCL) cruciate ligament with a fine needle. Visualisation of the structures appeared clearest from the posterior approach. From this approach 24 electrodes were inserted into the PCL and six into the ACL. From the anterior 16 were inserted into the ACL and six into the PCL. Twenty-one out of 24 electrodes were placed correctly into the PCL from the posterior view, and four out of six were placed correctly from the anterior view. From the posterior view four out of six were placed correctly in the ACL, and eight electrodes out of 16 were placed correctly from the anterior view. In conclusion, ultrasound guided insertion of electrodes from the posterior view can be performed with high accuracy into the PCL, whereas insertion into the ACL seems to be less accurate, probably because it is more difficult to visualise.

Synthesis of the stationary-phase sigma factor sigma s is positively regulated by ppGpp.

Strains of Escherichia coli which lack detectable guanosine 3',5'-bispyrophosphate (ppGpp) display a pleiotropic phenotype that in some respects resembles that of rpoS (katF) mutants. This led us to examine whether ppGpp is a positive regulator of sigma s synthesis. sigma s is a stationary-phase-specific sigma factor that is encoded by the rpoS gene. We found that a ppGpp-deficient strain is defective in sigma s synthesis as cells enter stationary phase in a rich medium, as judged by immunoblots. Under more-defined conditions we found that the stimulation of sigma s synthesis following glucose, phosphate, or amino acid starvation of wild-type strains is greatly reduced in a strain lacking ppGpp. The failure of ppGpp-deficient strains to synthesize sigma s in response to these starvation regimens could indicate a general defect in gene expression rather than a specific dependence of rpoS expression on ppGpp. We therefore tested the effect of artificially elevated ppGpp levels on sigma s synthesis either with mutations that impair ppGpp decay or by gratuitously inducing ppGpp synthesis with a Ptac::relA fusion. In both instances, we observed enhanced sigma s synthesis. Apparently, ppGpp can activate sigma s synthesis under conditions of nutrient sufficiency as well as during entry into stationary phase. This finding suggests that changes in ppGpp levels function both as a signal of imminent stationary phase and as a signal of perturbations in steady-state growth.

Overproduction and purification of sigma 32, the Escherichia coli heat shock transcription factor.

This paper reports the overproduction and the details of a rapid method to purify active sigma 32 that is free of core RNA polymerase enzyme. Maximal overproduction of sigma 32 in a T7 RNA polymerase-based expression system is achieved only in the presence of rifampicin. This 2-day procedure involves solubilizing inclusion bodies in Sarkosyl, removal of Sarkosyl by dialysis, and a single S-Sepharose column chromatography step. The final yield of sigma 32 is about 4.1 mg of approximately 95% purity from 1 g of wet weight cells.

In vitro functional characterization of overproduced Escherichia coli katF/rpoS gene product.

The katF/rpoS gene product (sigma s), a central regulator of stationary-phase gene expression in Escherichia coli, has been purified from an overproducing strain. sigma s was used as an immunogen for the production of monoclonal antibodies. Previous sequence analysis of sigma s strongly indicated homology to the sigma factor family. We show biochemically in this paper that sigma s is a sigma factor. This protein can bind to core RNA polymerase (E), and this binding can be competed effectively by the major E. coli transcription initiation factor, sigma 70. Immunopurified sigma s holoenzyme (E sigma s) transcribes the promoters of the bolAp1 gene and the xthA gene. Interestingly, both promoters can also be transcribed by sigma 70 holoenzyme (E sigma 70).

Interpretation of group behavior.

A Systematic Multiple Level Observation of Groups (SYMLOG) allows managers to graphically see how their perception of personal behavior compares with their colleagues' views of the same behavior. Through recognition and awareness, groups can change ineffective behavioral patterns.

[Ultrasonography of the rotator cuff of the shoulder]. / Ultralydundersøgelse af skulderens rotator-cuff.

Clinical examination for rotator cuff (RC) tear is rarely conclusive. Arthrography has been considered the standard diagnostic procedure for detection of rotator cuff tears. Ultrasonography (US) is less time-consuming and less expensive than arthrography and is, in addition, noninvasive. During recent years, many reports concerning US evaluation of RC tears have been published, but with different results about the diagnostic value of the examination. Recently, some authors have pointed out the ability of US in visualizing degenerative changes in the RC and that it is difficult to differentiate between tears and degenerative changes. By review of the English literature, we conclude that the experienced examiner with proper equipment can confidently differentiate between normal and pathological RC an that in quite a number of cases it is possible to differentiate tears from degenerative changes. In case of doubt, the examination should be supplemented by shoulder arthrography. In the hands of an experienced examiner with proper equipment US seems to be suitable as the initial imaging modality for evaluation of RC.

[Epitheloid sarcoma--an overlooked diagnosis?]. / Epiteloidt sarkom--en upåagtet diagnose?

Epitheloid sarcoma (ES) is a malignant soft-tissue tumour which occurs particularly on the hand and forearm in young adults. The tumour grows slowly and metastasizes relatively late but has, nevertheless, a considerable mortality and morbidity because it is frequently erroneously diagnosed, both clinically and pathologically, either as a benign tumour or as a reactive inflammatory process. Two typical cases with diagnostic delays of 12 and 15 years, respectively, which required amputations through the upper arm or forearm are described. Even although ES is a rare tumour form, clinicians and pathologists should be aware of this possibility as, in the early stages, the tumour can frequently be treated effectively and radically by wide local excision without amputation.

Epidemiology of meniscal lesions in the knee. 1,215 open operations in Copenhagen 1982-84.

In a suburban area of Copenhagen with approximately 620,000 inhabitants, all the openly operated on meniscal lesions of the knee joint between 1982 and 1984 inclusive were reviewed. The mean annual incidence of meniscal lesions per 10,000 inhabitants was 9.0 in males and 4.2 in females. The highest incidences were seen in the 3rd, 4th, and 5th decades of life. A higher frequency of trauma related to onset of symptoms was found among males (77 percent) than among females (64 percent). The bucket-handle lesion was the most frequent type of meniscal lesion in males (35 percent), whereas peripheral detachment was the most frequent in females (41 percent). A variable frequency of trauma inducing the lesions in males and females did not explain differences in type of meniscal lesion. Nineteen percent of our 1,215 patients had a partial meniscectomy, whereas 0.7 percent had meniscal repair.

The natural history of chondromalacia patellae. A 12-year follow-up.

A series of 45 patients with chondromalacia patellae demonstrated clinically and by arthrotomy, but without demonstrable biomechanical misalignment, was investigated retrospectively. Thirty-one of the patients had only been treated conservatively while partial chondrectomy was performed in 14. All of the patients underwent an intensive rehabilitation program. After 12 years the patients were interviewed. Eighty-one percent of the patients who were treated conservatively and 57 percent of the patients in whom partial chondrectomy was performed had mild or no pain and no other knee complaints. The results were better for patients aged 20 years or younger. Two patients received disability pensions on account of poor knee function and 6 had changed to less strenuous occupations. Restraint is advised as regards operative treatment of chondromalacia patellae.

Tibial plateau fractures. A comparison of conservative and surgical treatment.

We evaluated the long-term results of 109 tibial plateau fractures, 61 treated by skeletal traction and early knee movement and 48 treated by surgery, at an average follow-up of 70 months. The functional results were much the same, though meniscectomy had been performed in almost half of the surgical patients. Time in bed and duration of hospital stay were clearly shorter after surgery (p less than 0.0001). We concluded that conservative management is a valid alternative to surgery, but should probably be reserved for cases where operation is undesirable. Future studies should compare surgery without meniscectomy and conservative treatment using cast braces to reduce the time in traction.

Magnetic resonance imaging in the evaluation of sequelae after tibial plateau fractures.

Nine knees with persistent radiographic depression of the articular surface after tibial plateau fractures treated by traction and early knee motion were examined using magnetic resonance imaging (MRI). The MRI examinations demonstrated that the radiographic bone defects were filled up with different tissues, which we interpreted as cartilage, fibrous tissue and synovial plica. The type of tissue in the bone defect did not seem to affect the functional capabilities of the knee. However, a demonstrable thin cover of tissue, with the signal-intensity of cartilage, over the defect, seemed to be associated with an excellent functional result.

[Metastasizing squamous cell carcinoma in patients with psoriasis treated with immunosuppressive agents]. / Metastaserende spinocellulaert karcinom hos immunsupprimerede psoriasispatienter.

Immunosuppressive agents are imployed increasingly frequently in treatment of nonfatal disorders. We report two cases of metastasizing squamous cell carcinoma after treatment for psoriasis with methotrexate. Additional cases must be reported, but we consider that: 1) methotrexate medication, when possible, should be withdrawn when a localized squamous cell carcinoma is diagnosed and 2) when this diagnosis is confirmed the patient should be referred for radical treatment.