Long-term outcomes of very early treated infantile-onset Pompe disease with short-term steroid premedication: experiences from a nationwide newborn screening programme.

BACKGROUND: Starting enzyme replacement therapy (ERT) before severe irreversible muscular damage occurs is important in infantile-onset Pompe disease (IOPD). This long-term follow-up study demonstrates our diagnostic and treatment strategies for IOPD and compares our clinical outcomes with those of other medical centres. METHODS: In this long-term follow-up study, we analysed the outcomes of very early ERT with premedication hydrocortisone in patients with IOPD. Out of 1 228 539 infants screened between 1 January 2010 and 28 February 2021, 33 newborns had confirmed IOPD in Taipei Veterans General Hospital. Twenty-six were regularly treated and monitored at Taipei Veterans General Hospital. Echocardiographic parameters, biomarkers, IgG antibodies against alglucosidase alpha, pulmonary function variables and developmental status were all assessed regularly over an average follow-up duration of 6.18±3.14 years. We compared the long-term treatment outcomes of our patients with those of other research groups. RESULTS: The average age at ERT initiation was 9.75±3.17 days for patients with classic IOPD. The average of the latest antialglucosidase alpha IgG titre was 669.23±1159.23. All enrolled patients had normal heart sizes, motor milestones, cognitive function and pulmonary function that were near-normal to normal. Compared with patients in other studies, our patients had better outcomes in all aspects. CONCLUSION: Very early ERT using our rapid diagnostic and treatment strategy enabled our patients with IOPD to have better outcomes than patients in other medical centres.

The role of crystal packing on the optical response of trialkyltetrelethynyl acenes.

The electronic and optical responses of an organic semiconductor (OSC) are dictated by the chemistries of the molecular or polymer building blocks and how these chromophores pack in the solid state. Understanding the physicochemical nature of these responses is not only critical for determining the OSC performance for a particular application, but the UV/visible optical response may also be of potential use to determine aspects of the molecular-scale solid-state packing for crystal polymorphs or thin-film morphologies that are difficult to determine otherwise. To probe these relationships, we report the quantum-chemical investigation of a series of trialkyltetrelethynyl acenes (tetrel = silicon or germanium) that adopt the brickwork, slip-stack, or herringbone (HB) packing configurations; the π-conjugated backbones considered here are pentacene and anthradithiophene. For comparison, HB-packed (unsubstituted) pentacene is also included. Density functional theory and G0W0 (single-shot Green's function G and/or screened Coulomb function W) electronic band structures, G0W0-Bethe-Salpeter equation-derived optical spectra, polarized ϵ2 spectra, and distributions of both singlet and triplet exciton wave functions are reported. Configurational disorder is also considered. Furthermore, we evaluate the probability of singlet fission in these materials through energy conservation relationships.

Airway abnormalities and pulmonary complications in long-term treated late-onset Pompe disease: Diagnostic and interventional by flexible bronchoscopy.

This study evaluates the whole airway abnormalities of long-term treated late-onset Pompe disease (LOPD) patients, with interventions using the flexible bronchoscope (FB). As a retrospective study, we follow up with our five LOPD patients treated with Myozyme from 2012 to 2021 regularly, but with a focus on the whole airway abnormalities of these patients visualized through FB. The long-term clinical outcomes and relevant airway symptoms were assessed. Pulmonary function test and polysomnography were performed to evaluate the degree of respiratory compromise. All patients in the study had varying degrees of airway collapsibility, pulmonary complications, sleep apnea syndrome, and facial anomalies. Pulmonary function could preserve after Myozyme treatment, but potential deterioration thereafter. This is the first study that focuses on airway abnormalities and pulmonary complications in long-term treated LOPD patients using FB. Despite years of Myozyme treatment, we still observed airway abnormalities in these patients. In our series, the pulmonary complications seem more obvious than those observed in patients with infantile-onset Pompe disease, which might be related to the late diagnosis and treatment. We might recommend that FB could provide dynamic evaluation and interventions of airway abnormalities simultaneously. Early diagnosis of respiratory dysfunction is a critical prognostic factor of the long-term outcome of treated LOPD patients.

[Expression and role of Pim1 in cultured cortical neurons with oxygen-glucose deprivation/reoxygen injury].

OBJECTIVE: To study the expression and effect of Pim1 in primary cortical neurons after hypoxic-ischemic injury. METHODS: Cortical neurons were isolated from 1-day-old C57BL/6 mice and cultured in neurobasal medium. On the 8th day of neuron culture, cells were subjected to oxygen-glucose deprivation/reoxygen (OGD/R) treatment to mimic in vivo hypoxic injury of neurons. Briefly, medium were changed to DMEM medium, and cells were cultured in 1% O2 for 3 hours and then changed back to normal medium and conditions. Cells were collected at 0 hour, 6 hours, 12 hours and 24 hours after OGD/R. Primary neurons were transfected with Pim1 overexpression plasmid or mock plasmid, and then were exposed to normal conditions or OGD/R treatment. They were named as Pim1 group, control group, OGD/R group and OGD/R+Pim1 group respectively. Real-time PCR was used to detect Pim1 mRNA expression. Western blot was used to detect the protein expression of Pim1 and apoptotic related protein cleaved caspase 3 (CC3). TUNEL staining was used to detect cell apoptosis. RESULTS: Real-time PCR and Western blot results showed that Pim1 mRNA and protein were significantly decreased in neurons after OGD/R. They began to decrease at 0 hour after OGD/R, reached to the lowest at 12 hours after OGD/R, and remained at a lower level at 24 hours after OGD/R (P<0.01). Overexpression of Pim1 significantly upregulated the protein level of Pim1. Under OGD/R conditions, the CC3 expression and the apoptosis rate in cells of the Pim1 group were significantly lower than in un-transfected cells (P<0.01). CONCLUSIONS: Hypoxic-ischemic injury may decrease Pim1 expression in neurons. Overexpressed Pim1 may inhibit apoptosis induced by OGD/R.

Airway abnormalities in very early treated infantile-onset Pompe disease: A large-scale survey by flexible bronchoscopy.

Early enzyme replacement therapy (ERT) improve long-term outcomes in patients with infantile-onset Pompe disease (IOPD). Our cohort of patients with IOPD at Taipei Veterans General Hospital (TVGH) joined Taiwan Pompe newborn screening program from 2008, testing more than one million newborns until 2018. By 2010, we had established rapid diagnostic strategies. Now, the average age of ERT initiation starts at an average age of <10 days-old, the earliest group in the world. However, they still presented some airway problems. We present a retrospective study focused on airway abnormalities in these patients along 8 years of observation. Fifteen patients with IOPD, who received very early treatment at a mean age of 8.94 ± 3.75 days, underwent flexible bronchoscopy (FB) for dynamic assessment of the whole airway. Long-term clinical outcomes and relevant symptoms of the upper airway were assessed. All patients in the study had varying degrees of severity of upper airway abnormalities and speech disorders. The three oldest children (Age 94, 93, and 88 months, respectively) had poor movement of the vocal cords with reduced abduction and adduction and had silent aspiration of saliva through the glottis during respiration. This is the largest cohort study presented to date about airway abnormalities in very early treated patients with IOPD patients by FB. Despite very early treatment, we observed upper airway abnormalities in these IOPD patients. In IOPD, upper airway abnormalities seem inevitable over time. We suggest early and continuous monitoring for all IOPD patients, even with early and regular treatment.

[Effect of irisin on hypoxic-ischemic brain damage in neonatal rats].

OBJECTIVE: To study the effect and mechanism of action of irisin on hypoxic-ischemic brain damage in neonatal rats. METHODS: A total of 248 7-day-old Sprague-Dawley rats were randomly divided into a sham-operation group, a model group, and low- and high-dose irisin intervention groups (n=62 each). The rats in the model and irisin intervention groups were given hypoxic treatment after right common carotid artery ligation to establish a model of hypoxic-ischemic brain damage. Those in the sham-operation group were given the separation of the right common carotid artery without ligation or hypoxic treatment. The rats in the high- and low-dose irisin intervention groups were given intracerebroventricular injection of recombinant irisin polypeptide at a dose of 0.30 µg and 0.15 µg respectively. Those in the model and sham-operation groups were given the injection of an equal volume of PBS. The water maze test was used to compare neurological behaviors between groups. TTC staining, hematoxylin-eosin staining and TUNEL staining were used to observe histopathological changes of the brain. Western blot was used to measure the expression of the apoptosis-related molecules cleaved-caspase-3 (CC3), BCL-2 and BAX. RESULTS: Compared with the sham-operation group, the model group had a significant increase in latency time and a significant reduction in the number of platform crossings (P<0.05). Compared with the model group, the high-dose irisin intervention group had a significant reduction in latency time and a significant increase in the number of platform crossings (P<0.05). Compared with the sham-operation group, the model group had massive infarction in the right hemisphere, with significant increases in karyopyknosis and karyorrhexis. Compared with the model group, the high-dose irisin intervention group had a smaller infarct area of the right hemisphere, with reductions in karyopyknosis and karyorrhexis. The model group had a significantly higher apoptosis rate of cells in the right cerebral cortex and the hippocampus than the sham-operation group. The high-dose irisin intervention group had a significantly lower apoptosis rate than the model group (P<0.05). At 24 and 48 hours after modeling, the sham-operation group had a significantly lower level of CC3 than the model group (P<0.05). Compared with the model group, the high-dose irisin intervention group had a significantly lower level of CC3 and a significantly higher BCL-2/BAX ratio (P<0.05). The low-dose irisin intervention group had similar laboratory markers and histopathological changes of the brain to the model group. CONCLUSIONS: Irisin can alleviate hypoxic-ischemic brain damage in neonatal rats in a dose-dependent manner, possibly by reducing cell apoptosis in the cerebral cortex and the hippocampus.

Very Early Treatment for Infantile-Onset Pompe Disease Contributes to Better Outcomes.

OBJECTIVE: To evaluate whether very early treatment in our patients would result in better clinical outcomes and to compare these data with other infantile-onset Pompe disease (IOPD) cohort studies. METHODS: In this nationwide program, 669,797 newborns were screened for Pompe disease. We diagnosed IOPD in 14 of these newborns, and all were treated and followed in our hospital. RESULTS: After 2010, the mean age at first enzyme-replacement therapy (ERT) was 11.92 days. Our patients had better biological, physical, and developmental outcomes and lower anti-rh acid α-glucosidase antibodies after 2 years of treatment, even compared with one group that began ERT just 10 days later than our cohort. No patient had a hearing disorder or abnormal vision. The mean age for independent walking was 11.6 ± 1.3 months, the same age as normal children. CONCLUSIONS: ERT for patients with IOPD should be initiated as early as possible before irreversible damage occurs. Our results indicate that early identification of patients with IOPD allows for the very early initiation of ERT. Starting ERT even a few days earlier may lead to better patient outcomes.

Antioxidant properties of fruiting bodies, mycelia, and fermented products of the culinary-medicinal king oyster mushroom, Pleurotus eryngii (higher Basidiomycetes), with high ergothioneine content.

The culinary-medicinal king oyster mushroom Pleurotus eryngii is known to contain ergothioneine, and its products, including fruiting bodies, mycelia, and solid-state fermented products (adlay and buckwheat), were prepared to study their antioxidant properties. Fruiting bodies, regular and Hi-Ergo mycelia, and fermented products contained 2.05, 1.68, 5.76, 0.79-0.80 mg/g of ergothioneine, respectively. On the basis of the results obtained, P. eryngii products had effective antioxidant activity, reducing power, and scavenging ability on 1,1-diphenyl-2-picrylhydrazyl radicals and chelating ability on ferrous ions. Hi-Ergo mycelia was the most effective in the first 3 antioxidant properties in addition to its ergothioneine content. In addition, fruiting bodies were more effective in all antioxidant properties than regular mycelia. For ethanolic and hot water extracts from mycelia and fruiting bodies, the correlation coefficients between total phenol contents and each antioxidant attribute were 0.483-0.921. Overall, P. eryngii products with high amounts of ergothioneine could be used beneficially as a functional food.

Submerged cultivation of mycelium with high ergothioneine content from the culinary-medicinal king oyster mushroom Pleurotus eryngii (higher Basidiomycetes) and its composition.

The culinary-medicinal king oyster mushroom, Pleurotus eryngii, was used to produce mycelia with high ergothioneine content using a one-factor-at-a-time method. The optimal culture conditions for mycelia harvested at day 14 were 25°C, 10% inoculation rate, 2% glucose, 0.5% yeast extract, and no adjustment to the initial pH value. With histidine or amino acid mix added, biomasses and the ergothioneine content of mycelia were higher than those of the control. The ergothioneine content of mycelia harvested at days 16-20 were higher than that of mycelia harvested at day 14. In addition, the ergothioneine content of mycelia from the fermentor (5.84-5.76 mg/g) was much higher than that of mycelia from the shaken flask (4.93-5.04 mg/g). Mycelia with high ergothioneine content showed a profile of proximate composition similar to that of regular mycelia but lost its characteristic umami taste. Overall, mycelia high in ergothioneine could be prepared by optimal culture conditions, the addition of precursors, prolonged harvest, and aeration in the fermentor.

Preparation of culinary-medicinal king oyster mushroom Pleurotus eryngii-fermented products with high ergothioneine content and their taste quality.

Pleurotus eryngii (DC. : Fr.) Ouel. was used in solid-state fermentation to develop novel mushroom products with a high amount of ergothioneine. The correlation coefficients between ergothioneine content and biomass were 0.8878 and 0.9206 for fermented adlay and buckwheat, respectively. Using optimal conditions, Pleurotus-fermented adlay and buckwheat (PFA and PFB) with the ergothioneine contents of 795.5 and 786.1 mg/ kg, respectively, were prepared. However, mycelia contained the highest ergothioneine content of 1514.6 mg/kg. As a result of SSF by P. eryngii, PFA and PFB contained more taste components than adlay and buckwheat, as evidenced by higher contents of total sugars and polyols, total free amino acids and monosodium glutamate-like components, and total and flavor 5'-nucleotides. In addition, PFB and buckwheat showed comparable equivalent umami concentration (EUC) values, whereas PFA showed a higher EUC value than adlay. Overall, Pleurotus-fermented products with a high amount of ergothioneine will be a novel functional food.

Stepwise activity of ClpY (HslU) mutants in the processive degradation of Escherichia coli ClpYQ (HslUV) protease substrates.

In Escherichia coli, ClpYQ (HslUV) is a two-component ATP-dependent protease composed of ClpY (HslU), an ATPase with unfolding activity, and ClpQ (HslV), a peptidase. In the ClpYQ proteolytic complex, the hexameric rings of ClpY (HslU) are responsible for protein recognition, unfolding, and translocation into the proteolytic inner chamber of the dodecameric ClpQ (HslV). Each of the three domains, N, I, and C, in ClpY has its own distinct activity. The double loops (amino acids [aa] 137 to 150 and 175 to 209) in domain I of ClpY are necessary for initial recognition/tethering of natural substrates such as SulA, a cell division inhibitor protein. The highly conserved sequence GYVG (aa 90 to 93) pore I site, along with the GESSG pore II site (aa 265 to 269), contribute to the central pore of ClpY in domain N. These two central loops of ClpY are in the center of its hexameric ring in which the energy of ATP hydrolysis allows substrate translocation and then degradation by ClpQ. However, no data have been obtained to determine the effect of the central loops on substrate binding or as part of the processivity of the ClpYQ complex. Thus, we probed the features of ClpY important for substrate engagement and protease processivity via random PCR or site-specific mutagenesis. In yeast two-hybrid analysis and pulldown assays, using isolated ClpY mutants and the pore I or pore II site of ClpY, each was examined for its influence on the adjoining structural regions of the substrates. The pore I site is essential for the translocation of the engaged substrates. Our in vivo study of the ClpY mutants also revealed that an ATP-binding site in domain N, separate from its role in polypeptide (ClpY) oligomerization, is required for complex formation with ClpQ. Additionally, we found that the tyrosine residue at position 408 in ClpY is critical for stabilization of hexamer formation between subunits. Therefore, our studies suggest that stepwise activities of the ClpYQ protease are necessary to facilitate the processive degradation of its natural substrates.

Reversal of doxorubicin-resistance by multifunctional nanoparticles in MCF-7/ADR cells.

The efficacy of many chemotherapeutic agents is reduced in cells that have developed multiple drug resistance (MDR). To address this important problem, a biodegradable polymer was coupled to a photosensitizer and the resulting photosensitizer-nanoparticles were loaded with the chemotherapeutic agent doxorubicin. The combination of photosensitizer and chemotherapeutic agent had a synergistic action on a doxorubicin-resistant breast cancer MCF-7 cell line. To increase the effectiveness of this combination, d-alpha-tocopheryl poly(ethylene glycol) 1000 succinate (TPGS), an inhibitor of the multidrug transporter overproduced in these resistant cells, was added during the formation of the nanoparticles. The insertion of TPGS decreased the P-glycoprotein activity, increased the intracellular accumulation doxorubicin, and also increased the therapeutic efficacy of the resulting nanoparticles. Both TPGS and irradiation of the photoreactive nanoparticles caused doxorubicin to move from the cytoplasm to the nucleus. This combination of photodynamic activity in a powerful nanocarrier loaded with the chemotherapeutic agent doxorubicin can be used to deliver two types of cancer therapy simultaneously, and the addition of TPGS can further enhance the entry of doxorubicin into the nucleus. Therefore, this innovative delivery system can act as a potential nanomedicine for both drug-sensitive and drug-resistant cancer therapy.