Human iPSC-Derived 2D and 3D Platforms for Rapidly Assessing Developmental, Functional, and Terminal Toxicities in Neural Cells.

With increasing global health threats has come an urgent need to rapidly develop and deploy safe and effective therapies. A common practice to fast track clinical adoption of compounds for new indications is to repurpose already approved therapeutics; however, many compounds considered safe to a specific application or population may elicit undesirable side effects when the dosage, usage directives, and/or clinical context are changed. For example, progenitor and developing cells may have different susceptibilities than mature dormant cells, which may yet be different than mature active cells. Thus, in vitro test systems should reflect the cellular context of the native cell: developing, nascent, or functionally active. To that end, we have developed high-throughput, two- and three-dimensional human induced pluripotent stem cell (hiPSC)-derived neural screening platforms that reflect different neurodevelopmental stages. As a proof of concept, we implemented this in vitro human system to swiftly identify the potential neurotoxicity profiles of 29 therapeutic compounds that could be repurposed as anti-virals. Interestingly, many compounds displayed high toxicity on early-stage neural tissues but not on later stages. Compounds with the safest overall viability profiles were further evaluated for functional assessment in a high-throughput calcium flux assay. Of the 29 drugs tested, only four did not modulate or have other potentially toxic effects on the developing or mature neurospheroids across all the tested dosages. These results highlight the importance of employing human neural cultures at different stages of development to fully understand the neurotoxicity profile of potential therapeutics across normal ontogeny.

A case of 'blue skin' and 'dark urine'.

A 60-year-old female presented with a 20-year history of progressive dark bluish discoloration of skin and passage of dark colored urine, painful arthritis and a recent history of invasive ductal carcinoma of right breast. Skin biopsy revealed hyaline material which was Periodic-Acid-Schiff stain positive and Congo-red stain negative, urine analysis revealed dark urine with presence reducing substance and radio-imaging showed intervertebral ossification and joint ankyloses. The patient was managed symptomatically with physiotherapy and acetaminophen on as required basis. This image is reported as a classic case of Alkaptonuria with clinical, histopathological and radio-imaging findings and the presence of invasive ductal breast carcinoma in the same patient.

Pharmacological reversal of synaptic and network pathology in human MECP2-KO neurons and cortical organoids.

Duplication or deficiency of the X-linked MECP2 gene reliably produces profound neurodevelopmental impairment. MECP2 mutations are almost universally responsible for Rett syndrome (RTT), and particular mutations and cellular mosaicism of MECP2 may underlie the spectrum of RTT symptomatic severity. No clinically approved treatments for RTT are currently available, but human pluripotent stem cell technology offers a platform to identify neuropathology and test candidate therapeutics. Using a strategic series of increasingly complex human stem cell-derived technologies, including human neurons, MECP2-mosaic neurospheres to model RTT female brain mosaicism, and cortical organoids, we identified synaptic dysregulation downstream from knockout of MECP2 and screened select pharmacological compounds for their ability to treat this dysfunction. Two lead compounds, Nefiracetam and PHA 543613, specifically reversed MECP2-knockout cytologic neuropathology. The capacity of these compounds to reverse neuropathologic phenotypes and networks in human models supports clinical studies for neurodevelopmental disorders in which MeCP2 deficiency is the predominant etiology.

Functional and Mechanistic Neurotoxicity Profiling Using Human iPSC-Derived Neural 3D Cultures.

Neurological disorders affect millions of people worldwide and appear to be on the rise. Whereas the reason for this increase remains unknown, environmental factors are a suspected contributor. Hence, there is an urgent need to develop more complex, biologically relevant, and predictive in vitro assays to screen larger sets of compounds with the potential for neurotoxicity. Here, we employed a human induced pluripotent stem cell (iPSC)-based 3D neural platform composed of mature cortical neurons and astrocytes as a model for this purpose. The iPSC-derived human 3D cortical neuron/astrocyte co-cultures (3D neural cultures) present spontaneous synchronized, readily detectable calcium oscillations. This advanced neural platform was optimized for high-throughput screening in 384-well plates and displays highly consistent, functional performance across different wells and plates. Characterization of oscillation profiles in 3D neural cultures was performed through multi-parametric analysis that included the calcium oscillation rate and peak width, amplitude, and waveform irregularities. Cellular and mitochondrial toxicity were assessed by high-content imaging. For assay characterization, we used a set of neuromodulators with known mechanisms of action. We then explored the neurotoxic profile of a library of 87 compounds that included pharmaceutical drugs, pesticides, flame retardants, and other chemicals. Our results demonstrated that 57% of the tested compounds exhibited effects in the assay. The compounds were then ranked according to their effective concentrations based on in vitro activity. Our results show that a human iPSC-derived 3D neural culture assay platform is a promising biologically relevant tool to assess the neurotoxic potential of drugs and environmental toxicants.

Hepatitis B, interferon, and acne fulminans in a young girl.

Acne fulminans (AF) is a very rare severe form of acne seen in young males, characterized by a sudden and explosive onset of hemorrhagic pustules and ulceration on the trunk, systemic features in the form of fever, polyarthropathy, malaise, erythema nodosum and painful osteolytic bone involvement with leukocytosis, and an elevated erythrocyte sedimentation rate. Conventional treatment of AF includes corticosteroids or immunosuppressive agents for the initial phase followed by isotretinoin. Active hepatitis B infection with a high viral load precludes the administration of any immunosuppressive drugs. We present the case of an 18-year-old girl with a history of occasional acne who presented with AF of sudden onset following administration of interferon-alpha-2a for her recently detected hepatitis B infection. Management of hepatitis B was withheld in view of her general condition. The patient was managed with low dose isotretinoin with subsidence of lesions. AF in a young female precipitated by interferon and its management with isotretinoin in the presence of active hepatitis B infection make the case unique.

Congenital Erythropoietic Porphyria with Undescended Testis.

Hereditary porphyrias are a group of metabolic disorders of heme biosynthesis pathway that are characterized by acute neurovisceral symptoms, skin lesions, or both. Congenital erythropoietic porphyria (CEP) is an extremely rare disease with a mutation in the gene that codes for uroporphyrinogen III synthase leading to accumulation of porphyrin in different tissues and marked cutaneous photosensitivity. We report a case of CEP with infancy onset blistering, photosensitivity, red colored urine, and teeth along with scarring. Examination revealed an undescended testis of the left side. Mutation analysis revealed mutation in the uroporphyrinogen III synthase gene (UROS) resulting in c. 56 A > G (Tyr19Cys). The presence of undescended testis with a rare mutation in a case of CEP which itself is an extremely rare condition make the case interesting.

Incidence of amlodipine-induced gingival overgrowth in the rural population of Loni.

AIMS: Since the incidence of gingival overgrowth induced by amlodipine remains poorly defined, this study was carried out with an aim to determine the incidence. MATERIALS AND METHODS: Dental patients who received amlodipine (N = 115), for more than 3 months were studied to determine the drug-induced gingival overgrowth. Clinical diagnosis of drug-induced overgrowth was verified by disappearance or decreased severity of gingival overgrowth after withdrawal of the causative drug. RESULTS: The prevalence rate of amlodipine-induced gingival hyperplasia among experimental patients was 3.4%, while it was not observed among the control subjects. Oral examination revealed gingival overgrowth as a lobular or nodular enlargement on interdental papilla located in the anterior interproximal regions. CONCLUSIONS: In this study, there was a significant relationship between gingival inflammation resulting from dental plaque and drug dosage, and hyperplasia.

Expression of tumor necrosis factor α and its correlation with severity of oral submucous fibrosis: a case-control study.

OBJECTIVE: The objective of this case-control study was to evaluate the plasma tumor necrosis factor α (TNF-α) levels in patients with oral submucous fibrosis and to determine the relation of plasma TNF-α levels with the severity of oral submucous fibrosis. STUDY DESIGN: Blood samples were collected from 25 patients who were diagnosed with oral submucous fibrosis and from 25 age- and sex-matched control participants. The plasma was isolated by centrifugation of blood samples. The levels of plasma TNF-α were estimated using enzyme-linked immunosorbent assay. RESULTS: A total of 19 out of 25 patients had detectable plasma TNF-α levels ranging from 0.1 to 106.4 pg/mL (mean, 23.46 pg/mL), whereas only 12 out of 25 control participants had detectable plasma TNF-α levels ranging from 0.1 to 33.3 pg/mL (mean, 6.93 pg/mL). The difference between the TNF-α levels of patients and controls was statistically significant (P = .015) according to the Mann-Whitney test. CONCLUSIONS: Patients with oral submucous fibrosis had significantly increased TNF-α levels compared with controls.

A silent tumor of the gingiva: An unusual case report and surgical management with 1 year follow-up.

Neurofibroma is a benign tumor of the peripheral nerve sheath characterized by the proliferation of Schwann's cells, perineural cells, and endoneurial fibroblasts. Here, we present a case of unique variation in the observed characteristics of a neurofibroma, with no relation to neurofibromatosis (NF) type-1 or von Recklinghausen disease of the skin. A swelling was observed in the right maxillary gingiva in relation with 14-16 regions of a 25-year-old female patient. The lesion was smooth, with sessile base, painless, non-ulcerated, and with normal color. An excisional biopsy of the lesion was performed, and histologically the diagnosis was confirmed. The patient remains uneventful after 1 year of clinical follow-up. Oral cavity involvement by a solitary neurofibroma in patients with no other signs of NF is uncommon and the gingival involvement is very rare.