GelMA-Encapsulated hDPSCs and HUVECs for Dental Pulp Regeneration.

Pulpal revascularization is commonly used in the dental clinic to obtain apical closure of immature permanent teeth with thin dentinal walls. Although sometimes successful, stimulating bleeding from the periapical area of the tooth can be challenging and in turn may deleteriously affect tooth root maturation. Our objective here was to define reliable methods to regenerate pulp-like tissues in tooth root segments (RSs). G1 RSs were injected with human dental pulp stem cells (hDPSCs) and human umbilical vein endothelial cells (HUVECs) encapsulated in 5% gelatin methacrylate (GelMA) hydrogel. G2 RSs injected with acellular GelMA alone, and G3 empty RSs were used as controls. White mineral trioxide aggregate was used to seal one end of the tooth root segment, while the other was left open. Samples were cultured in vitro in osteogenic media (OM) for 13 d and then implanted subcutaneously in nude rats for 4 and 8 wk. At least 5 sample replicates were used for each experimental group. Analyses of harvested samples found that robust pulp-like tissues formed in G1, GelMA encapsulated hDPSC/HUVEC-filled RSs, and less cellularized host cell-derived pulp-like tissue was observed in the G2 acellular GelMA and G3 empty RS groups. Of importance, only the G1, hDPSC/HUVEC-encapsulated GelMA constructs formed pulp cells that attached to the inner dentin surface of the RS and infiltrated into the dentin tubules. Immunofluorescent (IF) histochemical analysis showed that GelMA supported hDPSC/HUVEC cell attachment and proliferation and also provided attachment for infiltrating host cells. Human cell-seeded GelMA hydrogels promoted the establishment of well-organized neovasculature formation. In contrast, acellular GelMA and empty RS constructs supported the formation of less organized host-derived vasculature formation. Together, these results identify GelMA hydrogel combined with hDPSC/HUVECs as a promising new clinically relevant pulpal revascularization treatment to regenerate human dental pulp tissues.

Whole genome sequencing and phylogenetic analysis of West Nile virus lineage 1 and lineage 2 from human cases of infection, Italy, August 2013.

A human outbreak of West Nile virus (WNV) infection caused by WNV lineage 2 is ongoing in northern Italy. Analysis of six WNV genome sequences obtained from clinical specimens demonstrated similarities with strains circulating in central Europe and Greece and the presence of unique amino acid changes that identify a new viral strain. In addition, WNV lineage 1 Livenza, responsible for a large outbreak in north-eastern Italy in 2012, was fully sequenced from a blood donor during this 2013 outbreak.

Human cases of West Nile Virus infection in north-eastern Italy, 15 June to 15 November 2010.

In 2010, for the third consecutive year, human cases of West Nile virus (WNV) infection, including three confirmed cases of neuroinvasive disease and three confirmed cases of West Nile fever, were identified in north-eastern Italy. While in 2008 and 2009 all human cases of WNV disease were recorded in the south of the Veneto region, cases of WNV disease in 2010 additionally occurred in two relatively small northern areas of Veneto, located outside those with WNV circulation in the previous years. WNV IgG antibody prevalence in blood donors resident in Veneto was estimated as ranging from 3.2 per 1,000 in areas not affected by cases of WNV disease to 33.3 per 1,000 in a highly affected area of the Rovigo province. No further autochthonous human cases of WNV disease were notified in Italy in 2010. The recurrence of human cases of WNV infection for the third consecutive year strongly suggests WNV has become endemic in north-eastern Italy.

Prevalence of IgM and IgG antibodies to West Nile virus among blood donors in an affected area of north-eastern Italy, summer 2009.

Following reports of West Nile neuroinvasive disease in the north-eastern area of Italy in 2009, all blood donations dating from the period between 1 August and 31 October 2009 in the Rovigo province of the Veneto region were routinely checked to exclude those with a positive nucleic acid test for West Nile virus (WNV). Only one of 5,726 blood donations was positive (17.5 per 100,000 donations; 95% confidence interval (CI): 0.4­97.3). In addition, a selection of 2,507 blood donations collected during the period from 20 July to 15 November 2009 were screened by ELISA for IgG and IgM antibodies against WNV. A positive result was received for 94 of them. The positive sera were further evaluated using immunofluorescence and plaque reduction neutralisation test (PRNT), in which only 17 sera were confirmed positive. This corresponds to a prevalence of 6.8 per 1,000 sera (95% CI: 4.0­10.9). In a case-control study that matched each of the 17 PRNT-positive sera with four negative sera with the same date of donation and same donation centre, we did not find a significant association with age and sex of the donor; donors who worked mainly outdoors were significantly more at risk to have a positive PRNT for WNV.

Genome sequence analysis of the first human West Nile virus isolated in Italy in 2009.

In 2009, six new human cases of West Nile neuroinvasive disease (WNND) were identified in Veneto region, following the six cases already reported in 2008. A human West Nile virus (WNV) isolate was obtained for the first time from an asymptomatic blood donor. Whole genome sequence of the human WNV isolate showed close phylogenetic relatedness to the Italy-1998-WNV strain and to other WNV strains recently isolated in Europe, with the new acquisition of the NS3-Thr249Pro mutation, a trait associated with avian virulence, increased virus transmission, and the occurrence of outbreaks in humans.

West Nile virus infection in Veneto region, Italy, 2008-2009.

We report here an update on human cases of West Nile virus (WNV) infection in Veneto region, northeastern Italy. In addition to two cases of WNV neuroinvasive disease notified through a surveillance programme started in September 2008, further four cases were retrospectively identified (in May 2009) by investigating patients with aseptic meningoencephalitis of unknown aetiology occurring in Veneto region in June-September 2008. All six patients had symptom onset in August-September 2008 and were resident in a wetland area close to the Po river delta in Rovigo province. Further five cases of asymptomatic WNV infection, including four residents of the same area in Rovigo, were identified in a seroprevalence study in farm workers from Veneto region. To date, no human cases have been notified in 2009.

Changes in the tonic activity of wrist extensor motor units induced by stimulating antagonistic group I afferents in humans.

The question as to whether the firing patterns of low- and high-threshold motor units in the extensor carpi radialis muscles are affected differently by group I afferents from the wrist flexors depending on the motor task being performed was investigated in six subjects. The motor units were voluntarily activated during a task consisting of either selectively contracting the wrist extensor muscles or co-activating the wrist and finger antagonist muscles by clenching the hand around a manipulandum. The motor units (n=40) were identified on the basis of their firing thresholds, their macro-potential areas, and their twitch contraction times. The effects on the motor-unit tonic activity of stimulating the wrist flexor afferents were assessed in terms of the changes in the firing probability, which were analysed after computing peri-stimulus time histograms using the cumulative sum procedure. Median nerve stimulation induced four main changes in the tonic firing pattern of the extensor motor units. An early, short-lasting increase in the firing probability (event E1) was found to occur in the high-threshold motor units, either in both tasks (6/13) or only during hand clenching (2/13). A short-latency decrease in the firing probability (event E2) was found to occur in all the motor units, the amount of which increased from the fast- to slowly contracting motor units, especially during hand clenching. A later decrease (event E3) followed by a large, late increase (event E4) in the tonic activity of the motor units was found to occur in all the motor units, without any task-dependent effects. All these various events were consistently observed in 12 pairs of motor units, each consisting of one slowly and one fast-contracting motor unit, which were tested simultaneously. These findings suggest that median nerve stimulation may selectively alter the tonic firing patterns of identified extensor motor units, depending on their functional characteristics (recruitment threshold, motor unit macro-potential area, contraction time) rather than on the excitatory drive to the motoneurone pool. The possible origins of these various events are discussed, and it is argued that, in the wrist extensor and flexor muscles that act as synergists during manipulatory finger movements and gripping tasks, the spinal pathways which assist the voluntary command may selectively modulate the firing patterns of identified motor units, to fit the requirements of the on-going motor task.

Successful VATS ligation of a large anomalous branch producing IMA steal syndrome after MIDCAB.

The occurrence of a flow "steal phenomenon" from a large branch of the internal mammary artery (IMA) is rare and its pathogenesis is still controversial. We describe a case of a patent large anomalous left IMA branch which produced recurrent angina 3 years post MIDCAB. Transcatheter coil obliteration of the vessel produced symptomatic relief. However, six months later, chest pains recurred in association with unwinding of the coil and recannulization of the collateral vessel. Successful ligation of the culprit branch using a VATS approach is presented.

Selective enhancement of motoneurone short-term synchrony during an attention-demanding task.

Recent experiments on monkeys suggest that attention-related changes in the synchronous activity of neurones occur in the motor cortex. In humans, the clinical data available suggest that the weak synchronization of the motor unit firing observed during voluntary contraction might reflect the activity of the motoneurone cortico-spinal afferents. The present study was therefore designed to investigate how the synchronous motor-unit activity might depend on the attention required in performing a motor task. Twenty-five motor unit pairs were tested in the right extensor carpi radialis muscles of five right-handed subjects, who were instructed to keep both motor units firing tonically while trying to maintain the extension force as constant as possible using visual feedback set either at low or high gain. Fifteen motor unit pairs (60%) showed a greater amount of synchronous activity, without any consistent changes in the motor unit firing rates, when the subjects were monitoring the force at high gain. In addition, the mean value and the steadiness of the force did not differ significantly between the two tasks. The amplitude of the motor units' contractile force extracted by spike-triggered averaging did not change consistently despite the slight increase in the synchronous impulse probability observed under the high-gain conditions. Changes in synchrony affected slowly contracting, low-threshold motor units as well as fast-contracting, high-threshold motor units. The most noteworthy finding was that these changes actually focused on a narrow component of the synchronous activity (within a period of less than 4 ms) in keeping with the short-term synchronization process. This suggests that common inputs, possibly of cortical origin, may have contributed more greatly to controlling the motoneurone firing pattern in the motor task which required the subjects to pay more attention.

Inhibition versus facilitation of the reflex responsiveness of identified wrist extensor motor units by antagonist flexor afferent inputs in humans.

The question of whether Ia reciprocal inhibition might depend on the motor task and on the type of motor unit activated was investigated in the human extensor carpi radialis muscles. Ia reciprocal inhibition induced by stimulating the median nerve (conditioning stimulation) was estimated by measuring the changes in the firing probability of 37 extensor motor units in response to the radial nerve stimulation (100 test stimuli) delivered 1 ms after the conditioning stimulation. Six subjects were asked to perform a task consisting of either selectively contracting their wrist extensor muscles or co-activating their wrist and finger antagonist muscles by clenching their hand around a manipulandum. In the control recordings (test stimulation alone), the mean response probability of the 37 motor units was found to be greater during hand clenching. The motor units were identified on the basis of their force thresholds, their macro-potentials, and their twitch contraction times. The data obtained in the control recordings were consistent with the size principle. In the recordings where the responses were conditioned by applying median nerve stimulation, the response probability of the motor units with low force thresholds, small macro-potential areas, and long twitch contraction times tended to decrease, in line with the presence of Ia reciprocal inhibition, whereas the response probability of the motor units with higher force thresholds, larger macro-potential areas, and shorter twitch contraction times tended to increase. The median nerve stimulation may therefore have altered the efficiency with which the extensor Ia inputs recruited the homonymous motoneurones in the pool. The flexor group I afferents activated while the median nerve was stimulated had inhibitory effects on the slow contracting motor units, and facilitatory effects mainly on the fast contracting motor units. Both of these effects were stronger during hand clenching, in which the numerous cutaneous receptors of the palm and fingertips are liable to be activated. Besides their own effects on the excitability of the various types of motor units, cutaneous inputs are known to potentiate the Ib interneurones. In addition, the effects of the conditioning stimulation were superimposed on the tonic activity of the Ia and Ib afferents from the flexor wrist and finger muscles. This may explain why both the inhibitory and facilitatory effects of the median nerve stimulation were enhanced during hand clenching.

Task dependence of Ia presynaptic inhibition in human wrist extensor muscles: a single motor unit study.

OBJECTIVE: Task-dependent changes in the Ia presynaptic inhibition generated by flexor group I afferents were investigated in 25 identified motor units (MUs) located in human extensor carpi radialis (ECR) muscles. METHODS: Seven subjects had to voluntarily contract their ECR muscles either alone during isometric wrist extension or concurrently with their wrist and finger flexor muscles while clenching their hand around a manipulandum. The MU reflex responses to the radial nerve stimulation (test stimulation) yielded narrow peaks in the post-stimulus time histograms (PSTH). The Ia presynaptic inhibition induced while stimulating the median nerve (conditioning stimulation) 20 and 40 ms before the radial nerve was assessed from the changes in the contents of the first 0.5 ms in the peaks. RESULTS: With both stimulation intervals, the Ia presynaptic inhibition, as assessed from the first 0.5 ms of the PSTH peaks, was consistently weaker during hand clenching. With both motor tasks, the Ia presynaptic inhibition was strongest at the 20 ms interval, in which it showed a downward gradient, working from slow to fast contracting MUs. With both intervals, the presynaptic inhibition was consistently weaker during hand clenching. The decrease in the Ia presynaptic inhibition observed at the 40 ms conditioning-test interval was less pronounced during wrist extension. CONCLUSION: It is suggested that the reason why Ia presynaptic inhibition was weaker during hand clenching may have been that this task involved numerous cutaneous inputs originating from the palm and finger tips. During gripping tasks, these cutaneous inputs may therefore contribute to adjusting the wrist stiffness by relieving the presynaptic inhibition.

Dual task-related changes in the synchronous activity of wrist extensor type-identified motor units in humans.

The coupling between the firings of 33 pairs of motor units tested in the extensor carpi radialis muscles was evaluated by cross-correlation analysis and compared during isometric wrist extension and hand clenching. A slightly greater amount of synchrony was observed during hand clenching (0.042 +/- 0.024 vs 0.035 +/- 0.017 synchronous impulses per trigger). This trend did not reach however the level of significance and the changes were actually found to be heterogeneous: in 15 out of the 33 pairs tested, synchronous activity with a narrow coupling (4.6 +/- 2.4 ms) consistent with short-term synchrony was greater during hand clenching whereas in nine other pairs, synchronous activity with a broader dispersion (9.0 +/- 4.5 ms) was reduced. These opposite changes could not be explained in terms of changes in the firing pattern of the motor units and were found instead to correlate with the motor units' biomechanical properties. Motor units with high recruitment thresholds and fast rising twitches showed predominantly an increase in synchrony; whereas the motor unit pairs with low recruitment thresholds and slow twitches showed either an increase or a decrease. The enhanced short-term synchrony suggests that common inputs distributed to motoneurones of all types were more effective during hand clenching whereas the decrease in weakly coupled synchronous activity suggests that other inputs synchronized at a pre-synaptic level and distributed more specifically to motor units recruited at low force levels were less effective. The possible origins of the inputs reflected in the dual changes are discussed in terms of the supra-spinal and peripheral pathways controlling the wrist extensor motoneurones during wrist extension and hand clenching.

Distribution of presynaptic inhibition on type-identified motoneurones in the extensor carpi radialis pool in man.

The question was addressed as to whether the magnitude of Ia presynaptic inhibition might depend on the type of motor unit activated during voluntary contraction in the wrist extensor muscles. For this purpose, we investigated the effects of applying electrical stimulation to the median nerve on the responses of 25 identified motor units to radial nerve stimulation delivered 20 ms after a conditioning stimulation. The reflex responses of the motor units yielded peaks in the post-stimulus time histograms with latencies compatible with monosynaptic activation. Although median nerve stimulation did not affect the motoneurone net excitatory drive assessed from the mean duration of the inter-spike interval, it led to a decrease in the contents of the first two 0.25 ms bins of the peak. This decrease may be consistent with the Ia presynaptic inhibition known to occur under these stimulation conditions. In the trials in which the median nerve was being stimulated, the finding that the response probability of the motor units, even in their monosynaptic components, tended to increase as their force threshold and their macro-potential area increased and as their twitch contraction time decreased suggests that the median nerve stimulation may have altered the efficiency with which the Ia inputs recruited the motoneurones in the pool. These effects were consistently observed in seven pairs of motor units each consisting of one slow and one fast contracting motor unit which were simultaneously tested, which suggests that the magnitude of the Ia presynaptic inhibition may depend on the type of motor unit tested rather than on the motoneurone pool excitatory drive. The present data suggest for the first time that in humans, the Ia presynaptic inhibition may show an upward gradient working from fast to slow contracting motor units which is able to compensate for the downward gradient in monosynaptic reflex excitation from 'slow' to 'fast' motor units. From a functional point of view, a weaker Ia presynaptic inhibition acting on the fast contracting motor units may contribute to improving the proprioceptive assistance to the wrist myotatic unit when the contraction force has to be increased.

Mechanical cutaneous stimulation alters Ia presynaptic inhibition in human wrist extensor muscles: a single motor unit study.

Reflex responses were evoked by radial nerve stimulation in 25 single motor units in the extensor carpi radialis muscles of seven subjects during voluntary isometric wrist extension. The responses consisted of narrow peaks in the post-stimulus time histograms with latencies compatible with monosynaptic activation. When the skin of the palm and finger tips was continuously swept using a soft rotating brush, the purely monosynaptic components of the motor unit responses, as assessed from the contents of the first two 0.25 ms bins of the peak, were found to increase. This increase did not affect the motoneurone net excitatory drive, as assessed by measuring the mean duration of the inter-spike intervals. The cutaneous inputs activated by the brush may have reduced the tonic presynaptic inhibition exerted on the Ia afferents homonymous to the extensor motor units tested. To further investigate whether Ia presynaptic inhibition was involved, the responses of the extensor motor units were conditioned by stimulating the median nerve 20 ms earlier, using a protocol which is known to induce Ia extensor presynaptic inhibition originating from flexor Ia afferents. The median nerve stimulation did not affect the motoneurone excitatory drive, but led to a decrease in the responses of the extensor motor units to the radial nerve stimulation, especially in the purely monosynaptic components. This decrease was consistent with the Ia presynaptic inhibition known to occur under these stimulation conditions. The cutaneous inputs activated by the brush were found to reduce the Ia presynaptic inhibition generated by the median nerve stimulation, without affecting the distribution of the Ia presynaptic inhibition among the various types of motor units tested. The present data suggest that cutaneous inputs from the palm and finger tips may relieve the Ia presynaptic inhibition exerted on the wrist extensor motor nuclei, and thus enhance the proprioceptive assistance to fit the specific requirements of the ongoing motor task.

Ia presynaptic inhibition in human wrist extensor muscles: effects of motor task and cutaneous afferent activity.

The task-dependence of the presynaptic inhibition of the muscle spindle primary afferents in human forearm muscles was studied, focusing in particular on the modulation associated with the co-contraction of antagonist muscles and the activation of cutaneous afferents. The changes known to affect the motoneuron proprioceptive assistance during antagonist muscle co-activation in human leg and arm muscles were compared. The evidence available so far that these changes might reflect changes in the presynaptic inhibition of the muscle spindle afferent is briefly reviewed. The possible reasons for changes in presynaptic inhibition during the antagonist muscle co-contraction are discussed. Some new experiments on the wrist extensor muscles are briefly described. The results showed that the changes in the Ia presynaptic inhibition occurring during the co-contraction of the wrist flexor and extensor muscles while the hand cutaneous receptors were being activated (the subject's hand was clenched around a manipulandum) could be mimicked by contracting the wrist extensor muscles alone while applying extraneous stimulation to the hand cutaneous receptors. It is concluded that besides the possible contribution of inputs generated by the co-contraction of antagonist muscles and by supraspinal pathways, cutaneous inputs may play a major role in modulating the proprioceptive assistance during manipulatory movements.

Sarcoma of the main pulmonary artery: an unusual etiology for recurrent pulmonary emboli.

We describe a case of primary pulmonary artery (PA) trunk spindle cell sarcoma in an 86 year old female presenting clinically with debilitating signs of recurrent pulmonary embolism. Further extensive work aroused suspicion for pulmonary artery malignancy. Palliative wide surgical resection, pulmonary artery tumor embolectomy and reconstruction of the proximal pulmonary artery and right ventricle outflow tract (RVOT) with bovine pericardial tissue were performed. She survived the procedure with an improved quality of life, but expired due to recurrence at 6 months postoperatively. Albeit uncommon, pulmonary artery sarcoma is nowadays a more frequently preoperatively diagnosed and surgically treated malignancy. With a modern low perioperative mortality, aggressive surgical resection remains as the single most effective modality for its treatment and can result in short term palliation in selected patients.

Proprioceptive control of wrist extensor motor units in humans: dependence on handedness.

The effectiveness of the monosynaptic proprioceptive assistance to the wrist extensor motoneurone activity was investigated during voluntary contraction in relation to the subjects' handedness. The reflex responses of 411 single motor units to homonymous tendon taps were recorded in the wrist extensor carpi radialis muscles in both arms of five right-handed and five left-handed subjects. In the right-handed subjects, the motor unit reflex responses were clearly lateralized in favour of their right arm, whereas no side-related differences were observed in the left-handed subjects, whatever the motor units' mechanical properties and firing rates. When the muscle spindle sensitivity was by-passed by electrically stimulating the primary afferents in both arms of three right-handed and three left-handed subjects, no side-related differences were observed in the Hoffmann reflex (H-reflex) amplitude in either of the two lateralization groups. The effectiveness of the primary afferent synapses on to the motoneurones therefore does not seem to depend on the subject's handedness. Without excluding the possibility of structural changes being involved at the periphery, the comparisons carried out on the data obtained using electrical vs mechanical stimulation suggest that the asymmetrical effectiveness of the proprioceptive assistance observed in favour of the right arm in the right-handed subjects might result from either the gamma or beta drive being more efficient. This asymmetry might result from the preferential use of the right hand in skilled movements. In a predominantly right-handed world, however, left-handed people might tend to develop the ability to use their right arm almost as skillfully as their preferred left arm, which could explain the symmetrical effectiveness of the proprioceptive assistance observed here in the left-handers' wrist extensor muscles.

Primary aorto-duodenal fistula secondary to infected abdominal aortic aneurysms: the role of local debridement and extra-anatomic bypass.

Gastrointestinal bleeding secondary to spontaneous rupture of an infected abdominal aortic aneurysm into the duodenum is a rare and highly lethal clinical occurrence, representing roughly a third of all primary aortoduodenal fistulas. Diagnosis is problematic due to the subtleties in the clinical presentation and course, and surgical treatment is usually delayed, representing a challenge even for the experienced vascular surgeon. The overall mortality is over 30% and the operative approaches are still controversial. Two cases of ruptured infrarenal aortic aneurysms complicated with aortoduodenal fistula were recently treated at our institution. Bacterial aortitis was documented by arterial wall cultures positive for Klebsiella and Salmonella species respectively. The clinical courses and outcomes of the two patients (one survivor ) treated with retroperitoneal debridement and extra-anatomic bypass and a review of the modern surgical treatment are herein described.

Pulmonary resection for malignancy in the elderly: is age still a risk factor?

OBJECTIVE: There is an increasing number of elderly patients presenting with potentially-resectable lung malignancy. The objective of this study is to evaluate the modern perioperative morbidity and mortality in patients undergoing oncologic lung resection and to analyse the trend over a 26-year period in our experience. METHODS: Between 1971 and 1996, 1506 patients underwent lung resection for malignancy. We reviewed the 30-day perioperative risk in a group of 385 (25.6%) patients aged 70 years and older operated on for intended cure of lung malignancy. Operations included 293 (77%) lobectomies, 24 pneumonectomies (6%), 16 bilobectomies (4%) and 52 wedge or segmental resections (13%). The pathology was bronchogenic carcinoma in 89% and metastasis or other tumours in 11% of patients. We compared the 30-day perioperative risk between the elderly group (age 70 or greater) and a cohort of 180 patients (control) 69 years and younger. RESULTS: The mortality for all resections in elderly group was 4.2% (16/385) and was 1.6% for the control group. Mortality in the octogenarian group was 2.8%. Female gender correlated with a decreased risk of death, with only two of 16 deaths in females (P < 0.005). Overall morbidity was higher in the study than in control patients (34% vs. 25%, n.s.), although major morbidity was similar in both groups (13.2% vs. 13%). Abnormal pulmonary-function testing and positive cardiac history did not correlate with increase overall or specific risk. Pneumonectomy carried a higher risk for death, with three of 24 deceased (12.5%; P < 0.05). Changes in outcome were analysed over two time periods: the mortality in the early period (1971-1982), 11.1% (8/72), was significantly elevated above the control group, while mortality in the modern period (1983-1994) was not, with a rate of 2.6% (8/313). CONCLUSIONS: In our series, mortality associated with operative treatment for lung malignancy in the elderly declined, so age alone no longer appears to be a risk factor. Age remains a risk factor for overall, but not major, morbidity. Pneumonectomy should undertaken cautiously in this age group. Based on this data, functional elderly patients should not be denied curative lung resection based on age alone.