Anticancer Activity of Aaptos suberitoides on Glioblastoma Multiforme Cell Line.

OBJECTIVE: Glioblastoma Multiforme (GBM) poses a significant challenge due to its high aggressiveness and unfavorable prognosis, with existing treatments demonstrating limited efficacy in prolonging survival rates. This study aimed to assess the anticancer properties of Aaptos suberitoides extracts and fraction on the U87 cell line, serving as a representative model for GBM. METHODS: U87 cells were treated with ethanol extracts derived from Aaptos suberitoides, specifically two extracts (OAA-1 and OAA-2) and one ethyl acetate fraction (EA) isolated from specimens collected on Pramuka Island and Tinjil Island. The evaluation encompased microscopic observation and MTT assay to determine the IC50. Subsequently, antiproliferative effects were investigated through apoptosis and cell cycle assays. RESULTS: The extract demonstrated cytotoxic activity against U87 cells, with OAA-1 and OAA-2 exhibiting IC50 values of 35.78 µg/mL and 25.38 µg/mL, respectively. OAA-1 notably induced apoptosis at 50 µg/mL and induced cell cycle arrest. On other hand, OAA-2, while also inducing apoptosis significantly, had a lesser impact on cell cycle arrest. In contrast, EA induced significant apoptosis at a concentration of 100 µg/mL. CONCLUSION: The ethanol extracts and the ethyl acetate fraction of Aaptos suberitoides emerged as a promising candidate for Glioblastoma Multiforme cancer therapy, showing potential in inhibiting cell proliferation and inducing apoptosis.

Low-cost and open-source three-dimensional (3D) printing in neurosurgery: A pilot experiment using direct drive modification to produce multi-material neuroanatomical models.

BACKGROUND: 3-dimensional (3D) printing carries a genuine potential for pre-operative planning in neurosurgery. Entry-level 3D printers offer practicality in low resource settings, but are often limited by the range of filament materials as well as the capability of open-source segmentation software. OBJECTIVE: We intended to demonstrate that 3D printing of neuroanatomical structures is possible using an entry-level 3D printer equipped with the direct drive (DD) modification, which supported flexible filaments, with the models segmented using an open-source software. METHODS: A DD system was installed onto the Ender 3 Pro printer. An attempt to print neurosurgical models using a low-cost 3D printer was conducted, where four patient-based neuroanatomical models were printed: skull base-vasculature, skull base-tumour, cervical spine, and ventricular system. The results were discussed and compared to similar endeavours in past literature. RESULTS: Although DD installation was challenging and led to vibration and longer print time, which ultimately warranted an inferior printing speed, DD system enabled the printing with thermoplastic polyurethane (TPU), a versatile elastomer; in addition to producing equal amount of detail to those printed with high-end printers and advanced image segmentation software. Fitting the frame well, changing infill type, and avoiding warping and stringing will improve print quality with the DD system. CONCLUSION: 3D printing with entry-level 3D printers equipped with DD system has been proven to be a reliable way of accurately reproducing patient-specific neuroanatomical constructs. Follow-up studies are necessary to implement 3D printing for neurosurgical planning in low-resource settings.

A Rarely Occurring Spinal Fracture Precipitated by Generalized Spasms of Tetanus Patient with Spondylitis Tuberculosis.

Background: The occurrence of spinal fracture due to tetanus nowadays is extremely rare, as compared to the 1950s, since the widely available anti-tetanus and antispasmodic therapy. The spinal fracture in tetanus patients is usually reported in higher thoracic vertebrae, previously with a rate as high as 57.5%. Spondylitis is the most common form of skeletal tuberculosis (TB) and can cause a spinal fracture. In Indonesia, tetanus is still reported, while tuberculosis is still endemic; however, co-infection of both diseases is rarely reported. Case Presentation: A 36-year-old male was brought to our hospital with jaw stiffness, accompanied by fever. A history of dental cavities was present, and 5 days prior, he experienced a fishing hook wound on his right index finger. There was no history of TB. Physical examination showed meningismus, 2 cm trismus, abdominal spasm, opisthotonus, and spontaneous muscle spasms, without dysautonomia. In the third week of hospitalization, while his tetanus condition improved, he complained of weakness in both legs. A thorough history taking revealed a history of backache for 3 years. A wedge-shaped fracture on his 11th and 12th thoracic vertebrae was observed on radiographic examination. A spinal TB diagnosis was made, and treatment was started. He refused to get spinal surgery, then went home with 4 out of 5 motor strength scale. After three months, he returned to his routine activity as a food hawker with no motor deficits. Conclusion: Tetanus spinal fracture is extremely rare nowadays; a thorough history of spinal problems/medication is compulsory for anticipation. This patient's spinal fracture was deemed due to a preexisting TB spinal infection that was precipitated by prolonged continuous tetanic spasm due to general tetanus.

Brain MRI Volumetry Analysis in an Indonesian Family of SCA 3 Patients: A Case-Based Study.

Introduction: Spinocerebellar ataxia type-3 (SCA3) is an adult-onset autosomal dominant neurodegenerative disease. It is caused by expanding of CAG repeat in ATXN3 gene that later on would affect brain structures. This brain changes could be evaluated using brain MRI volumetric. However, findings across published brain volumetric studies have been inconsistent. Here, we report MRI brain volumetric analysis in a family of SCA 3 patients, which included pre-symptomatic and symptomatic patients. Methodology: The study included affected and unaffected members from a large six-generation family of SCA 3, genetically confirmed using PolyQ/CAG repeat expansion analysis, Sanger sequencing, and PCR. Clinical evaluation was performed using Scale for the Assessment and Rating of Ataxia (SARA). Subjects' brains were scanned using 3.0-T MRI with a 3D T1 BRAVO sequence. Evaluations were performed by 2 independent neuroradiologists. An automated volumetric analysis was performed using FreeSurfer and CERES (for the cerebellum). Result: We evaluated 7 subjects from this SCA3 family, including 3 subjects with SCA3 and 4 unaffected subjects. The volumetric evaluation revealed smaller brain volumes (p < 0.05) in the corpus callosum, cerebellar volume of lobules I-II, lobule IV, lobule VIIB and lobule IX; and in cerebellar gray matter volume of lobule IV, and VIIIA; in the pathologic/expanded CAG repeat group (SCA3). Conclusion: Brain MRI volumetry of SCA3 subjects showed smaller brain volumes in multiple brain regions including the corpus callosum and gray matter volumes of several cerebellar lobules.

Clinical and genetic profile in index patients with spinocerebellar ataxia type 3 in Indonesia: case report.

Spinocerebellar ataxia (SCA) is an autosomal dominant hereditary disease with progressive course, and no causal therapy. Diagnostics are still challenging, due to facility and protocols, and so as in Indonesia. As a national referral center, Dr. Hasan Sadikin Central General Hospital has received a lot of patients from all over Indonesia, particularly from Western Java. Study related to SCA (including clinical and genetic profile) is still limited in Indonesia. We identified index patients from three families with ataxia, hence intend to determine their clinical and genetic pattern. The hereditary pattern is autosomal dominant. Scale for the assessment and rating of ataxia (SARA) shows mild and moderate ataxia. Inventory of non-ataxia signs (INAS) scores of the patients were 3, 5 and 6. Montreal cognitive assessment-Indonesian version (MOCA-INA) shows only one patient has mild cognitive impairment, despite young age. Barthel index shows 1 subject has moderate dependency. Mutation in Ataxin3 polyQ repeats shows pathologically long CAG repeats, 72,10; 72,10; and 72,23 respectively in mutant and wild type allele. We diagnosed the index patients with spinocerebellar ataxia type 3. This study is the first case series study in Indonesia. The hereditary pattern is clearly shown as an autosomal dominant ataxia. The clinical and genetic profile was varied, and the symptom is progressive and deteriorates overtime, including wide based gait, speech problem, motor and sensor complaint, and cognitive decline complaint. Despite the same polyQ stretch length, the onset and clinical characteristics of patients are diverse.

The Ethanol Extract of Marine Sponge Aaptos suberitoides Suppress Cell Viability, Cell Proliferation and Cell Migration in HER2-Positive Breast Cancer Cell Line.

OBJECTIVE: This study aimed to investigate the cytotoxicity, anti-proliferation and anti-migration effect of the ethanol extract of Aaptos suberitoides on trastuzumab-resistant HER2+ breast cancer cell line. METHODS: Aaptos suberitoides was collected from Tinjil Island, Banten, Indonesia, and was processed with maceration and ethanol extraction. HCC-1954 cells were treated with the ethanol extract and then followed by 3- [4, 5-dimethylthiazol-2-yl] -2.5 diphenyl tetrazolium bromide (MTT) assay to assess cytotoxicity, clonogenic assay and three-dimensional (3D) spheroid assay to evaluate anti-proliferative effect in two-dimensional and 3D model, respectively, and wound healing assay to determine anti-cell migration effect. Four parametric regression was used to analyse the IC50. RESULTS: This study revealed that the ethanol extract of Aaptos suberitoides suppressed cell viability in correlation with cell death induction. The IC50 values of the ethanol extract of Aaptos suberitoides using MTT assay and clonogenic assay were 12.0 ppm and 4.36 ppm, respectively. The extract demonstrated an inhibition effect on spheroid growth. In low concentration, the extract of Aaptos suberitoides inhibited cell migration. Furthermore, MS analysis showed that the most abundant compounds in this extract has molecular weight m/z 229.81 [M+H]+. CONCLUSION: This study revealed that the ethanol extract of Aaptos suberitoides demonstrates cytotoxicity, anti-proliferation and anti-migration effect as well as inhibition effect on three-dimensional spheroid growth in trastuzumab-resistant HER2+ breast cancer cell line.
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Bioactive Compounds in the Ethanol Extract of Marine Sponge Stylissa carteri Demonstrates Potential Anti-Cancer Activity in Breast Cancer Cells

Objective: Despite advanced treatment options available, drug resistance develops in breast cancer (BC) patients requiring novel effective drugs. Stylissa carteri, a marine sponge predominantly living in Indonesia territories, has not been extensively studied as anti-cancer. Therefore, this study targeted to assess the anti-tumor activity of the ethanol extract of S. carteri in BC cells. Methods: S. carteri was collected from Pramuka Island, at Kepulauan Seribu National Park, Jakarta, Indonesia and extracted using ethanol. Different BC cells including MDA MB 231, MDA MB 468, SKBR3, HCC-1954 and MCF-7 cells were treated with this extract for cytotoxic analysis using MTT assay. Spheroid growth assay and apoptosis assay were conducted in HCC-1954 cells. In addition, cell migration analysis and synergistic activity with doxorubicin or paclitaxel were conducted in MDA MB 231 cells. This extract was subjected also for GC-MS analysis. Results: The results show that ethanol extract of S. carteri demonstrated a cytotoxic activity in BC cells. The IC50 of this extract was lower 15 µg/ml in MDA MB 231, MDA MB 468, SKBR3, and HCC-1954 cells. Moreover, this extract inhibited spheroids growth and induced apoptosis in HCC-1954 cells. It inhibited cell migration and demonstrated a synergistic activity with doxorubicin or paclitaxel on triggering cell death in MDA MB 231 cells. Furthermore, GC-MS analysis indicated that this extract contained 1,2-Benzenediol, Dibutyl phthalate and 9,12-Octadecadienoic acid, ethyl ester. Conclusion: Our preliminary data indicate a potential anti-tumor activity of ethanol extract of S. carteri in breast cancer cells.

Fusion of Human Fetal Mesenchymal Stem Cells with "Degenerating" Cerebellar Neurons in Spinocerebellar Ataxia Type 1 Model Mice.

Mesenchymal stem cells (MSCs) migrate to damaged tissues, where they participate in tissue repair. Human fetal MSCs (hfMSCs), compared with adult MSCs, have higher proliferation rates, a greater differentiation capacity and longer telomeres with reduced senescence. Therefore, transplantation of quality controlled hfMSCs is a promising therapeutic intervention. Previous studies have shown that intravenous or intracortical injections of MSCs result in the emergence of binucleated cerebellar Purkinje cells (PCs) containing an MSC-derived marker protein in mice, thus suggesting a fusion event. However, transdifferentiation of MSCs into PCs or transfer of a marker protein from an MSC to a PC cannot be ruled out. In this study, we unequivocally demonstrated the fusion of hfMSCs with murine PCs through a tetracycline-regulated (Tet-off) system with or without a Cre-dependent genetic inversion switch (flip-excision; FLEx). In the FLEx-Tet system, we performed intra-cerebellar injection of viral vectors expressing tetracycline transactivator (tTA) and Cre recombinase into either non-symptomatic (4-week-old) or clearly symptomatic (6-8-month-old) spinocerebellar ataxia type 1 (SCA1) mice. Then, the mice received an injection of 50,000 genetically engineered hfMSCs that expressed GFP only in the presence of Cre recombinase and tTA. We observed a significant emergence of GFP-expressing PCs and interneurons in symptomatic, but not non-symptomatic, SCA1 mice 2 weeks after the MSC injection. These results, together with the results obtained using age-matched wild-type mice, led us to conclude that hfMSCs have the potential to preferentially fuse with degenerating PCs and interneurons but not with healthy neurons.

A CDC42EP4/septin-based perisynaptic glial scaffold facilitates glutamate clearance.

The small GTPase-effector proteins CDC42EP1-5/BORG1-5 interact reciprocally with CDC42 or the septin cytoskeleton. Here we show that, in the cerebellum, CDC42EP4 is exclusively expressed in Bergmann glia and localizes beneath specific membrane domains enwrapping dendritic spines of Purkinje cells. CDC42EP4 forms complexes with septin hetero-oligomers, which interact with a subset of glutamate transporter GLAST/EAAT1. In Cdc42ep4(-/-) mice, GLAST is dissociated from septins and is delocalized away from the parallel fibre-Purkinje cell synapses. The excitatory postsynaptic current exhibits a protracted decay time constant, reduced sensitivity to a competitive inhibitor of the AMPA-type glutamate receptors (γDGG) and excessive baseline inward current in response to a subthreshold dose of a nonselective inhibitor of the glutamate transporters/EAAT1-5 (DL-TBOA). Insufficient glutamate-buffering/clearance capacity in these mice manifests as motor coordination/learning defects, which are aggravated with subthreshold DL-TBOA. We propose that the CDC42EP4/septin-based glial scaffold facilitates perisynaptic localization of GLAST and optimizes the efficiency of glutamate-buffering and clearance.

Mutant ataxin-3 with an abnormally expanded polyglutamine chain disrupts dendritic development and metabotropic glutamate receptor signaling in mouse cerebellar Purkinje cells.

Spinocerebellar ataxia type 3 (SCA3) is caused by the abnormal expansion of CAG repeats within the ataxin-3 gene. Previously, we generated transgenic mice (SCA3 mice) that express a truncated form of ataxin-3 containing abnormally expanded CAG repeats specifically in cerebellar Purkinje cells (PCs). Here, we further characterize these SCA3 mice. Whole-cell patch-clamp analysis of PCs from advanced-stage SCA3 mice revealed a significant decrease in membrane capacitance due to poor dendritic arborization and the complete absence of metabotropic glutamate receptor subtype1 (mGluR1)-mediated retrograde suppression of synaptic transmission at parallel fiber terminals, with an overall preservation of AMPA receptor-mediated fast synaptic transmission. Because these cerebellar phenotypes are reminiscent of retinoic acid receptor-related orphan receptor α (RORα)-defective staggerer mice, we examined the levels of RORα in the SCA3 mouse cerebellum by immunohistochemistry and found a marked reduction of RORα in the nuclei of SCA3 mouse PCs. To confirm that the defects in SCA3 mice were caused by postnatal deposition of mutant ataxin-3 in PCs, not by genome disruption via transgene insertion, we tried to reduce the accumulation of mutant ataxin-3 in developing PCs by viral vector-mediated expression of CRAG, a molecule that facilitates the degradation of stress proteins. Concomitant with the removal of mutant ataxin-3, CRAG-expressing PCs had greater numbers of differentiated dendrites compared to non-transduced PCs and exhibited retrograde suppression of synaptic transmission following mGluR1 activation. These results suggest that postnatal nuclear accumulation of mutant ataxin-3 disrupts dendritic differentiation and mGluR-signaling in SCA3 mouse PCs, and this disruption may be caused by a defect in a RORα-driven transcription pathway.

Distinct transduction profiles in the CNS via three injection routes of AAV9 and the application to generation of a neurodegenerative mouse model.

Using single-stranded adeno-associated virus serotype 9 (ssAAV9) vectors containing the neuron-specific synapsin-I promoter, we examined whether different administration routes (direct cerebellar cortical (DC), intrathecal (IT) and intravenous (IV) injections) could elicit specific transduction profiles in the CNS. The DC injection route robustly and exclusively transduced the whole cerebellum, whereas the IT injection route primarily transduced the cerebellar lobules 9 and 10 close to the injection site and the spinal cord. An IV injection in neonatal mice weakly and homogenously transduced broad CNS areas. In the cerebellar cortex, the DC and IT injection routes transduced all neuron types, whereas the IV injection route primarily transduced Purkinje cells. To verify the usefulness of this method, we generated a mouse model of spinocerebellar ataxia type 1 (SCA1). Mice that received a DC injection of the ssAAV9 vector expressing mutant ATXN1, a protein responsible for SCA1, showed the intranuclear aggregation of mutant ATXN1 in Purkinje cells, significant atrophy of the Purkinje cell dendrites and progressive motor deficits, which are characteristics of SCA1. Thus, ssAAV9-mediated transduction areas, levels, and cell types change depending on the route of injection. Moreover, this approach can be used for the generation of different mouse models of CNS/neurodegenerative diseases.