Is epidural analgesia non-inferior to intrathecal fentanyl as initiation for neuraxial analgesia in early non-spontaneous labour?

BACKGROUND AND AIM: Intrathecal fentanyl, using the combined spinal-epidural (CSE) technique, provides rapid analgesia during early labour. Because of the technique's more complex and invasive nature, as its replacement we assessed the use of epidural analgesia in primiparous parturients with induced labour. The study was registered at www. CLINICALTRIALS: gov (NCT04645823). The aim was to compare the efficacy, duration of analgesia and maternal satisfaction. The primary outcome was the difference in pain visual analogue scale (VAS) between the interventions at 20 min after the analgesia administration. METHODS: Sixty volunteering parturients were randomly allocated in 1:1 ratio to receive either intrathecal fentanyl 20 µg or epidural analgesia (fentanyl 100 µg and lidocaine 80 mg). Contraction pain and maternal satisfaction were assessed by 0-100 mm VAS for 30 min, respectively. Foetal heart rate abnormalities, the time to first epidural dose and the incidence of pruritus were recorded. Non-inferiority margin for mean (95% CI) VAS after epidural analgesia was set at 20 mm above the VAS value for intrathecal fentanyl at 20 min. RESULTS: The contraction pain VAS fell from (median [interquartile range, IQR]) 82 (14) to 13 (20) mm and 76 (17) to 12 (27) mm in 20 min following the intrathecal fentanyl and epidural analgesia, respectively. The absolute mean difference (epidural-intrathecal fentanyl) in the VAS values was 3.3(-0.06 to 6.66) mm indicating non-inferiority. The median time to reach VAS <30 mm was 10 min in both groups. The duration until request for supplemental analgesia was 82(69-95) and 91(75-106) min after intrathecal fentanyl and epidural analgesia, respectively. The difference for the duration (epidural-intrathecal fentanyl) was 9 (6-12) min and for satisfaction-VAS 0.3 (-3.0 to 3.7) mm. There were no differences between the groups in the incidence of foetal heart rate abnormalities, while pruritus was more common after intrathecal fentanyl. CONCLUSION: After 20 min, epidural analgesia by lidocaine and fentanyl was within the non-inferior threshold compared with intrathecal fentanyl in efficacy. The duration of action was not shorter than that of intrathecal fentanyl and maternal satisfaction was also similar.

Labour analgesia by single shot spinal for any parturient?-A retrospective one-year single centre audit.

BACKGROUND: Single shot spinal (SSS) provides effective analgesia for multiparous parturients during advanced labour. Its utility in early labour or primiparous parturients may be limited by the insufficient duration of action. Regardless, SSS may offer a reasonable labour analgesia option in certain clinical scenarios. In this retrospective study, we analyse the failure rate of SSS analgesia by assessing pain after the SSS and by determining the need for additional analgesic interventions in primiparous or early-stage multiparous parturients compared to multiparous parturients in advanced labour (cervix ≥6 cm). METHODS: Following institutional ethical board approval, the patient files of all parturients receiving SSS analgesia during a 12-month period in a single centre were analysed for any recorded notes regarding recurrent pain or subsequent analgesia interventions (a new SSS, epidural, pudendal or paracervical bloc) as a marker for insufficient analgesia. RESULTS: A total of 88 primiparous and 447 multiparous parturients (cervix <6 cm: N = 131; cervix ≥6 cm: N = 316) received SSS analgesia. The odds ratio for the insufficient duration of analgesia was 1.94 (1.08-3.48) in primiparous and 2.08 (1.25-3.46) in early-stage multiparous parturients compared to advanced multiparous labour (p < .01). Primiparous and early-stage multiparous parturients were also 2.20 (1.15-4.20) and 2.61 (1.50-4.55) times more likely, respectively, to receive new peripheral and/or neuraxial analgesic interventions during delivery (p < .01). CONCLUSIONS: SSS appears to provide adequate labour analgesia for the majority of parturients in whom it is used, including nulliparous and early-stage multiparous parturients. It remains a reasonable option in certain clinical scenarios, including resource-limited settings where epidural analgesia is unavailable.

A Family With A20 Haploinsufficiency Presenting With Novel Clinical Manifestations and Challenges for Treatment.

BACKGROUND: Tumor necrosis factor α-induced protein 3 gene (TNFAIP3, also called A20) haploinsufficiency (HA20) leads to autoinflammation and autoimmunity. We have recently shown that a p.(Lys91*) mutation in A20 disrupts nuclear factor κB signaling, impairs protein-protein interactions of A20, and leads to inflammasome activation. METHODS: We now describe the clinical presentations and drug responses in a family with HA20 p.(Lys91*) mutation, consistent with our previously reported diverse immunological and functional findings. RESULTS: We report for the first time that inflammasome-mediated autoinflammatory lung reaction caused by HA20 can be treated with interleukin 1 antagonist anakinra. We also describe severe anemia related to HA20 successfully treated with mycophenolate. In addition, HA20 p.(Lys91*) was found to associate with autoimmune thyroid disease, juvenile idiopathic arthritis, psoriasis, liver disease, and immunodeficiency presenting with specific antibody deficiency and genital papillomatosis. CONCLUSIONS: We conclude that HA20 may lead to combination of inflammation, immunodeficiency, and autoimmunity. The condition may present with variable and unpredictable symptoms with atypical treatment responses.

Spinal and epidural sufentanil and fentanyl in early labour.

BACKGROUND AND AIM: The analgesic effect on labour pain of either spinal or epidural sufentanil or fentanyl was tested in a total of 80 primiparous parturients at an early phase of the delivery. The aim of the study was to compare the level of analgesia achieved within 20 minutes. METHODS: The parturients were randomly assigned to groups receiving either spinal sufentanil (5 µg), epidural sufentanil (20 µg), spinal fentanyl (20 µg) or epidural fentanyl (100 µg), whereafter the parturients were monitored for reported pain during contraction and side effects for 30 minutes. The primary outcome was the level of analgesia achieved within 20 minutes, while the secondary outcome was the time until the administration of the first epidural bolus. RESULTS: At baseline, the mean maximum pain VAS was 86 (84-89) mm. At 20 minutes after spinal sufentanil, epidural sufentanil, spinal fentanyl or epidural fentanyl, the maximum VAS was 19 (7-31), 45 (32-59), 25 (10-39) or 52 (40-63) mm, respectively (P < .01 spin vs epid groups). There were no differences in efficacy between spinal or epidural sufentanil and fentanyl. The mean (95% CI) time to the activation of epidural analgesia was 151 (111-192), 130 (93-168), 177 (121-234) and 112 (80-143) minutes after spinal sufentanil, epidural sufentanil, spinal fentanyl and epidural fentanyl, respectively. CONCLUSIONS: In terms of a reduction of VAS score at 20 minutes, epidural sufentanil or fentanyl provide 63% and 60% of the analgesic effect of the corresponding spinal analgesia. Epidural sufentanil or fentanyl could be used in situations in which spinal/CSE administration is not possible or desired.

A retrospective study of pre-operative fasting times prior to elective or emergency cesarean birth in a large maternity hospital: Lessons to be learned to minimize the fasting time.

INTRODUCTION: When managing elective and emergency cesarean births in the same operating room, unpredictable variations in the start times of the cesareans can prolong fasting periods. METHODS: The fasting times were retrospectively analyzed on 279 consecutive cesarean births at Helsinki University Women's Hospital, Finland, during January-February 2023. The fasting times were compared between the urgency groups and for elective cesareans according to their scheduled order on the operation list. The primary outcome was the difference in the fasting times for food and drink, while the secondary outcome was fasting for both food >12 h and fluids >4 h. The fasting times were compared by one-way ANOVA and chi-squared test, respectively. Dichotomous data are presented as unadjusted odds ratios (OR with 95% CI). RESULTS: Increasing urgency was associated with shorter fasting times. Fasting times for elective cesareans increased with the scheduled order on the daily list. The mean fasting periods (SD) increased from 10.55 h (SD=1.57) to 14.75 h (SD=2.02) from the first to the third cesarean of the day (p<0.01). The unadjusted odds ratio (95% CI) for fasting of the scheduled cesareans to exceed 12 h for solid foods and 4 h for clear fluids was 6.53 (95% CI: 2.67-15.9, p<0.001), for the third and fourth cesareans compared to the first two cesareans of the day. CONCLUSIONS: When elective and emergency cesareans are performed by the same team, the woman undergoing the third elective surgery of the day should be advised to have breakfast before 5 a.m. at home. While waiting for the operation, a carbohydrate drink should be offered to limit the fast.

Intrathecal atipamezole augments the antinociceptive effect of morphine in rats.

BACKGROUND: Opioid analgesics are effective in the treatment of chronic pain, but they have serious adverse effects such as development of tolerance and dependence. Adrenergic α(2) agonists and µ-opioid receptor agonists show synergistic potentiation and cross-tolerance in spinal analgesia, whereas α(2)-adrenergic antagonists have shown pronociceptive effects. However, at ultralow doses, spinal α(2)-adrenergic antagonists have been reported to paradoxically enhance opioid antinociception. New data have suggested a functional µ-opioid-α(2)-adrenoceptor complex, which may help in interpreting the paradoxical effect of the α(2)-adrenergic antagonists. In the present study we assessed the effects of low doses of atipamezole, a nonselective α(2)-adrenergic antagonist, on both systemic and spinal morphine antinociception and tolerance. METHODS: Antinociception was assessed in male Sprague-Dawley rats using hotplate, tail-flick, and paw pressure tests. Spinal or systemic opioid tolerance was induced for 4 days. The effects of both intrathecal and subcutaneous atipamezole on acute morphine-induced antinociception and established morphine tolerance were studied. RESULTS: Systemic or spinal atipamezole itself did not produce antinociception at the doses studied (subcutaneous 0.03, 0.3, 3 µg/kg or intrathecal 0.1, 1, 10 ng). The combined administration of spinal morphine and 1 ng of atipamezole increased the antinociceptive effect of acute spinal morphine 30 minutes after the administration of test drugs in the tail-flick test. Furthermore, 10 ng of intrathecal atipamezole attenuated established morphine tolerance 30 minutes after the administration of test drugs in the tail-flick test. However, subcutaneous atipamezole had no significant effect on systemic morphine antinociception, and it did not attenuate morphine tolerance. CONCLUSIONS: Spinal coadministration of low doses of atipamezole augmented the antinociceptive effect of morphine in naïve and tolerant rats. Heterodimerization of µ-opioid- and α(2A)-adrenoceptors with consequent changes in function and interaction could explain these results. This also suggests an interesting explanation for the variability in opioid response and tolerance in patients experiencing stress or having an increased noradrenergic tone due to other causes, e.g., drugs.

Successful pregnancy and cesarean delivery in a tetraplegic, home-invasively-mechanically-ventilated patient - case report.

INTRODUCTION: Pregnancies are rare in patients with severely disabilitating spinal cord injuries (SCI) but increasing alongside social awareness concerning reproductive equality. Physicians should be aware of several potential complications during pregnancy and delivery, particularly autonomic dysreflexia. CASE PRESENTATION: We report a successful pregnancy of a 32-year-old woman with a severe SCI at the C2 level (C1-4 ASIA Impairment Scale grade A) and total dependency on home invasive mechanical ventilation (HIMV), an extremely rare treatment. An elective cesarean section was chosen as the delivery mode at 34 + 0 weeks of gestation. Both the mother and the child recovered well. DISCUSSION: Severe spinal cord injury and dependency on mechanical ventilation are not absolute contraindications for pregnancy. With careful planning, pregnancy is possible also for patients with the most severe forms of SCI. Adequate pain relief during cesarean delivery is required despite complete spinal cord injury in order to avoid excessive hemodynamic responses and spinal reflexes. A multidisciplinary team is needed to ensure safe pregnancy and delivery of these high-risk pregnancies.

Proposal of a novel Artificial Intelligence Distribution Service platform for healthcare.

In this paper, we focus on presenting a novel AI-based service platform proposal called AIDI (Artificial Intelligence Distribution Interface for healthcare). AIDI proposal is based on our earlier research work in which we evaluated AI-based healthcare services which have been used successfully in practice among healthcare service providers. We have also used our systematic review about AI-based healthcare services benefits in various healthcare sectors. This novel AIDI proposal contains services for health assessment, healthcare evaluation, and cognitive assistant which can be used by researchers, healthcare service provides, clinicians, and consumers. AIDI integrates multiple health databases and data lakes with AI service providers and open access AI algorithms. It also gives healthcare service providers open access to state-of-the-art AI-based diagnosis and analysis services. This paper provides a description of AIDI platform, how it could be developed, what can become obstacles in the development, and how the platform can provide benefits to healthcare when it will be operational in the future.

Recovery after enhanced versus conventional care laparoscopic hysterectomy performed in the afternoon: A randomized controlled trial.

OBJECTIVE: To compare enhanced recovery after surgery (ERAS) and conventional care (CC) protocols on outcomes of laparoscopic hysterectomy (LH) performed in the afternoon. METHODS: A single-center randomized controlled trial was conducted on 120 women undergoing LH who were randomly divided into the intervention group (IG; n=60) and control group (CG; n=60). Women in the IG were treated according to the ERAS protocol and those in the CG according to the CC protocol. The primary outcome was the length of hospitalization. Secondary outcomes were postoperative opioid use, postoperative pain and emesis, complications, operative bleeding, and time. RESULTS: More women discharged during 24 hours in the IG than in the CG (88% vs 55%, P<0.001). The time to actual discharge (19 vs 22 hours, P<0.001) and ready-to-discharge time (15 vs 21 hours, P<0.001) were shorter and the use of oxycodone was lower (0 mg [0-0 vs 2.5 mg [0-10], P<0.001) in the IG than in the CG, respectively. Otherwise, no other significant differences between the groups were observed. The follow-up time was one month. CONCLUSION: The ERAS protocol reduces hospital stay and decreases the use of opioids with no impairment in surgical outcome of LH. ClinicalTrials.gov: NCT03828981.

Persistent susceptibility of cathepsin B to irreversible inhibition by nitroxyl (HNO) in the presence of endogenous nitric oxide.

Nitrosation of enzyme regulatory cysteines is one of the key posttranslational modification mechanisms of enzyme function. Frequently such modifications are readily reversible; however, cysteine proteases, such as cathepsin B, have been shown to be covalently and permanently inactivated by nitroxyl (HNO), the one-electron reduction product of NO. Owing to the high reactivity of HNO with NO, endogenous NO production could provide direct protection for the less reactive protein cysteines by scavenging HNO. Additionally, endogenous cellular production of NO could rescue enzyme function by protective nitrosation of cysteines prior to exposure to HNO. Thus, we studied the effect of endogenous NO production, induced by LPS or IFN-gamma, on inhibition of cysteine protease cathepsin B in RAW macrophages. Both LPS and IFN-gamma induce iNOS with generation of nitrate up to 9 muM in the media after a 24-h stimulation, while native RAW 264.7 macrophages neither express iNOS nor generate nitrate. After the 24-h stimulation, the HNO-releasing Angeli's salt (0-316 microM) caused dose-dependent and DTT-irreversible loss of cathepsin B activity, and induction of iNOS activity did not protect the enzyme. The lack of protection was also verified in an in vitro setup, where papain, a close structural analogue of cathepsin B, was inhibited by Angeli's salt (2.7 microM) in the presence of the NO donor DEA/NO (0-316 microM). This clearly showed that a high molar excess of DEA/NO (EC(50) 406 microM) is needed to protect papain from the DTT-irreversible covalent modification by HNO. Our results provide first evidence on a cellular level for the remarkably high sensitivity of active-site cysteines in cysteine proteases for modification by HNO.

Spatial and temporal distribution of laminins in permanent focal ischemic brain damage of the adult rat.

Laminins are extracellular matrix glycoproteins with multiple functions in the central nervous system, including maintenance of the blood-brain barrier. Because ischemic brain damage results in rapid degradation of extracellular matrix, we used immunocytochemistry on rat central nervous system after permanent focal ischemia to identify laminins involved in pathophysiology of stroke. At 24 hr after stroke, laminin-1 is transiently expressed by neurons inside the ischemic core, but from 2-3 days to 28 days it is expressed only in basement membrane structures. During the first 24 hr, alpha1, alpha5, beta1, and gamma1 laminins are transiently expressed in neurons within the ischemic core as an acute reaction of the brain to ischemia. Rapid induction of gamma1 laminin but no other laminin in reactive astrocytes surrounding the ischemic core is clear at 24 hr, and importantly, expression of gamma1 laminin in astrocytes surrounding the ischemic core intensifies during the first days and persists up to 28 days after stroke. At 2-3 days, gamma1 laminin immunoreactive barrier of reactive astrocytes is already fully formed, isolating the ischemic area from the healthy brain. Similar to gamma1 laminin, its KDI domain localizes in reactive astrocytes isolating the ischemic core. Results indicate that gamma1 laminin and its KDI domain are rapidly induced in glial cells after stroke and their expression persists, forming a molecular barrier between the healthy and the damaged brain. Thus, gamma1 laminin is involved in pathology of stroke and is likely to serve a protective function, considering its potent neuroprotective role after spinal cord injury and in neurodegenerative disorders.

Haploinsufficiency of A20 impairs protein-protein interactome and leads into caspase-8-dependent enhancement of NLRP3 inflammasome activation.

OBJECTIVES: TNFAIP3 encodes A20 that negatively regulates nuclear factor kappa light chain enhancer of activated B cells (NF-κB), the major transcription factor coordinating inflammatory gene expression. TNFAIP3 polymorphisms have been linked with a spectrum of inflammatory and autoimmune diseases and, recently, loss-of-function mutations in A20 were found to cause a novel inflammatory disease 'haploinsufficiency of A20' (HA20). Here we describe a family with HA20 caused by a novel TNFAIP3 loss-of-function mutation and elucidate the upstream molecular mechanisms linking HA20 to dysregulation of NF-κB and the related inflammasome pathway. METHODS: NF-κB activation was studied in a mutation-expressing cell line using luciferase reporter assay. Physical and close-proximity protein-protein interactions of wild-type and TNFAIP3 p.(Lys91*) mutant A20 were analysed using mass spectrometry. NF-κB -dependent transcription, cytokine secretion and inflammasome activation were compared in immune cells of the HA20 patients and control subjects. RESULTS: The protein-protein interactome of p.(Lys91*) mutant A20 was severely impaired, including interactions with proteins regulating NF-κB activation, DNA repair responses and the NLR family pyrin domain containing 3 (NLRP3) inflammasome. The p.(Lys91*) mutant A20 failed to suppress NF-κB signalling, which led to increased NF-κB -dependent proinflammatory cytokine transcription. Functional experiments in the HA20 patients' immune cells uncovered a novel caspase-8-dependent mechanism of NLRP3 inflammasome hyperresponsiveness that mediated the excessive secretion of interleukin-1ß and interleukin-18. CONCLUSIONS: The current findings significantly deepen our understanding of the molecular mechanisms underlying HA20 and other diseases associated with reduced A20 expression or function, paving the way for future therapeutic targeting of the pathway.

Increased catechol-O-methyltransferase activity and protein expression in OX-42-positive cells in the substantia nigra after lipopolysaccharide microinfusion.

Activated microglial cells are found in the substantia nigra and the striatum of Parkinson's disease patients. These cells have been shown to express catechol-O-methyltransferase activity which may increase during pathological conditions. Lipopolysaccharides are potent activators of microglial cells. After paranigral lipopolysaccharide infusion to rats we observed intense microglial activation around the lesion area followed by a delayed injury in nigrostriatal pathway in 2 weeks. Simultaneously, catechol-O-methyltransferase activity in the substantia nigra was gradually increased up to 213%. In the Western blot the amount of soluble COMT and membrane bound COMT proteins were increased by 255% and 86%, respectively. Increased catechol-O-methyltransferase immunoreactivity was located primarily into the activated microglial cells in the lesion area. Interestingly, catechol-O-methyltransferase and OX-42 stained also intensively microglia/macrophage-like cells which surrounded the adjacent blood vessels. Inhibition of catechol-O-methyltransferase activity by tolcapone or entacapone did not increase lipopolysaccharide-induced neurotoxicity. We conclude that catechol-O-methyltransferase activity and protein expression were increased in the substantia nigra after inflammation induced by lipopolysaccharides. These changes in glial and perivascular catechol-O-methyltransferase activity may have clinical relevance for Parkinson's disease drug treatment due to increased metabolism of levodopa in the brain.

Cathepsin B is a differentiation-resistant target for nitroxyl (HNO) in THP-1 monocyte/macrophages.

We previously showed that the one-electron reduction product of nitric oxide (NO), nitroxyl (HNO), irreversibly inhibits the proteolytic activity of the model cysteine protease papain. This result led us to investigate the differential effects of the nitrogen oxides, such as nitroxyl (HNO), NO, and in situ-generated peroxynitrite on cysteine modification-sensitive cellular proteolytic enzymes. We used Angeli's salt, diethylaminenonoate (DEA/NO), and 3-morpholinosydnoniminehydrochloride (SIN-1), as donors of HNO, NO, and peroxynitrite, respectively. In this study we evaluated their inhibitory activities on the lysosomal mammalian papain homologue cathepsin B and on the cytosolic 26S proteasome in THP-1 monocyte/macrophages after LPS activation or TPA differentiation. HNO-generating Angeli's salt caused a concentration-dependent (62 +/- 4% at 316 muM) inhibition of the 26S proteasome activity, resulting in accumulation of protein-bound polyubiquitinylated proteins in LPS-activated cells, whereas neither DEA/NO nor SIN-1 showed any effect. Angeli's salt, but not DEA/NO or SIN-1, also caused (94 +/- 2% at 316 muM) inhibition of lysosomal cathepsin B activity in LPS-activated cells. Induction of macrophage differentiation did not significantly alter the inhibitory effect of HNO on lysosomal cathepsin B activity, but protected the proteasome from HNO-induced inhibition. The protection awarded by macrophage differentiation was associated with induction of the GSH synthesis rate-limiting enzyme gamma-glutamylcysteine synthetase, as well as with increased intracellular GSH. In conclusion, HNO abrogates both lysosomal and cytosolic proteolysis in THP-1 cells. Macrophage differentiation, associated with upregulation of antioxidant defenses such as increased cellular GSH, does not protect the lysosomal cysteine protease cathepsin B from inhibition.

Nitric oxide-related species-induced protein oxidation: reversible, irreversible, and protective effects on enzyme function of papain.

Protein oxidation, irreversible modification, and inactivation may play key roles in various neurodegenerative disorders. Therefore, we studied the effects of the potentially in vivo occurring nitric oxide-related species on two different markers of protein oxidation: protein carbonyl generation on bovine serum albumine (BSA) and loss of activity of a cysteine-dependent protease, papain, in vitro by using Angeli's salt, papanonoate, SIN-1, and S-nitrosoglutathione (GSNO) as donors of nitroxyl, nitric oxide, peroxynitrite, and nitrosonium ions, respectively. Angeli's salt, SIN-1, and papanonoate (0-1000 microM) all generated a concentration-dependent increase in carbonyl formation on BSA (107, 60, and 45%, respectively). GSNO did not affect carbonyl formation. Papain was inhibited by Angeli's salt, SIN-1, papanonoate, and GSNO with IC50 values of 0.62, 2.3, 54, and 80 microM, respectively. Angeli's salt (3.16 microM)-induced papain inactivation was only partially reversible, while the effects of GSNO (316 microM) and papanonoate (316 microM) were reversible upon addition of excess DTT. The Angeli's salt-mediated DTT-irreversible inhibition of papain was prevented by GSNO or papanonoate pretreatment, hypothetically through mixed disulfide formation or S-nitrosylation of the catalytically critical thiol group of papain. These results, for the first time, compare the generation of carbonyls in proteins by Angeli's salt, papanonoate, and SIN-1. Furthermore, these results suggest that S-nitrosothiols may have a novel function in protecting critical thiols from irreversible oxidative damage.

Angeli's salt and spinal motor neuron injury.

UNLABELLED: Nitroxyl anion or its conjugate acid (NO-/HNO) and nitric oxide (NO) may both have pro-oxidative and cytotoxic properties. Superoxide dismutase (SOD) enzyme has been shown to convert reversibly HNO to NO. Mutations found in the SOD enzyme in some familial amyotrophic lateral sclerosis (ALS) patients affect redox properties of the SOD enzyme in a manner, which may affect the equilibrium between NO and HNO. Therefore, we studied the effects of HNO releasing compound, Angeli's salt (AS), on both motor and sensory functions after intrathecal administration in the lumbar spinal cord of a male rat. These functions were measured by rotarod, spontaneous activity, paw- and tail-flick tests. In addition, we compared the effect of AS to NO releasing papanonoate, old AS solution and sulphononoate in the motor performance test. The effect of intrathecal delivery of AS on the markers of the spinal cord injury and oxidative/nitrosative stress were further studied. RESULTS: Freshly prepared AS (5 or 10 micromol), but not papanonoate, caused a marked decrease in the rotarod performance 3-7 days after the intrathecal administration. The peak motor deficiency was noted 3 days after AS (5 micromol) delivery. Old, degraded, AS solution and nitrous oxide releasing sulphononoate did not decrease motor performance in the rotarod test. AS did not affect the sensory stimulus evoked responses as measured by the paw-flick and tail-flick tests. Immunohistological examination revealed that AS caused injury related changes in the expression of glial fibrillary acidic protein (GFAP), fibroblast growth factor (FGF-2) and laminins in the spinal cord. Moreover, AS increased nitrotyrosine immunoreactivity in the spinal motor neurons. Therefore, we conclude that AS, but not NO releasing papanonoate, causes motor neuron injury but does not affect the function of sensory nerves in behavioural tests.

Selective overexpression of gamma1 laminin in astrocytes in amyotrophic lateral sclerosis indicates an involvement in ALS pathology.

Our earlier studies indicate that the KDI tripeptide of gamma1 laminin reverts paralysis and protects adult rat CNS from excitotoxicity of glutamate and from oxidative stress. Here we show that gamma1 laminin is selectively overexpressed in reactive astrocytes of the amyotrophic lateral sclerosis (ALS) spinal cord, with both gray and white matter astrocytes overexpressing gamma1 laminin. Intensely gamma1 laminin-positive, aggressive-looking reactive astrocytes of the lateral columns of both cervical and thoracic spinal cord surround the lateral ventral horns and roots and extend into the area of the lateral corticospinal tract. In the cervical ALS spinal cord, large numbers of strongly gamma1 laminin-immunoreactive astrocytes are also present in the dorsal columns of the ascending sensory pathways. No other laminin or any other ALS-associated protein localizes in this manner. This unique distribution of gamma1 laminin-immunoreactive astrocytes in the ALS white matter together with our recent results on the efficacy of the KDI domain as a neuronal protector strongly suggest that gamma1 laminin may be expressed by astrocytes of the ALS spinal cord as a protective measure intended to aid neuronal survival. Further comparative studies on ALS spinal cord tissues and those of the animal models of ALS are needed to clarify the specific role of gamma1 laminin and its KDI domain in ALS and its putative interactions with the additional ALS-associated factors, such as excitotoxicity, oxidative stress, and neurofilament accumulation. Most importantly, further studies are urgently needed to test the potential of the KDI tripeptide as a therapeutic treatment for ALS.

KDI tripeptide of gamma1 laminin protects rat dopaminergic neurons from 6-OHDA induced toxicity.

Our previous studies indicate that the KDI (Lys-Asp-Ile) tripeptide of gamma1 laminin protects central neurons from mechanical trauma and excitotoxicity. At least part of the neuroprotective effect of the KDI tripeptide may be mediated by its inhibitory function on ionotropic glutamate receptors. We studied the protective effect of the KDI tripeptide against 6-hydroxy-dopamine (6-OHDA) induced neurotoxicity in a rat experimental model of Parkinson's disease (PD). We found that a single unilateral injection of the KDI tripeptide into the substantia nigra before an injection of 6-OHDA protected the dopaminergic neurons from the neurotoxicity of 6-OHDA. Compared to rats treated with 6-OHDA alone, the KDI + 6-OHDA-treated substantia nigra was relatively intact with large numbers of dopaminergic neurons present at the injection side. In the rats treated with 6-OHDA alone, no dopaminergic neurons were detected, and the substantia nigra-area at the injection side was filled with blood-containing cavities. Quantification of the rescue effect of the KDI tripeptide indicated that, in animals receiving KDI before 6-OHDA, 33% of tyrosine hydroxylase-positive dopaminergic neurons of the substantia nigra were present as compared to the contralateral non-injected side. In animals receiving 6-OHDA alone, only 1.4% of the tyrosine hydroxylase expressing dopaminergic neurons could be verified. If this much protection were achieved in humans, it would be sufficient to diminish or greatly alleviate the clinical symptoms of PD. We propose that the KDI tripeptide or its derivatives might offer a neuroprotective biological alternative for treatment of PD.

Regeneration of adult rat spinal cord is promoted by the soluble KDI domain of gamma1 laminin.

Regeneration in the central nervous system (CNS) of adult mammals is hampered by formation of a glial scar and by proteins released from the myelin sheaths of injured neuronal pathways. Our recent data indicate that the KDI (Lys-Asp-Ile) domain of gamma1 laminin neutralizes both glial- and myelin-derived inhibitory signals and promotes survival and neurite outgrowth of cultured human spinal cord neurons. We show that after complete transection of the adult rat spinal cord, animals receiving onsite infusion of the KDI domain via osmotic mini-pumps recover and are able to sustain their body weights and walk with their hindlimbs. Animals treated with placebo suffer from irreversible hindlimb paralysis. Microscopic and molecular analyses of the spinal cords indicate that the KDI domain reduces tissue damage at the lesion site and enables neurite outgrowth through the injured area to effect functional recovery of the initially paralyzed animals. That the KDI domain enhances regeneration of acute spinal cord injuries in the adult rat suggests that it may be used to promote regeneration of spinal cord injuries in humans.