Non-mitotic proliferation of malignant cancer cells revealed through live-cell imaging of primary and cell-line cultures.

INTRODUCTION: Anti-mitosis has been a key strategy of anti-cancer therapies, targeting at a fundamental property of cancer cells, their non-controllable proliferation due to overactive mitotic divisions. For improved anti-cancer therapies, it is important to find out whether cancer cells can proliferate independent of mitosis and become resistant to anti-mitotic agents. RESULTS: In this study, live-cell imaging was applied to both primary-cultures of tumor cells, and immortalized cancer cell lines, to detect aberrant proliferations. Cells isolated from various malignant tumors, such as Grade-III hemangiopericytoma, atypical meningioma, and metastatic brain tumor exhibit distinct cellular behaviors, including amoeboid sequestration, tailing, tunneling, nucleic DNA leakage, as well as prokaryote-like division such as binary fission and budding-shedding, which are collectively referred to and reported as 'non-mitotic proliferation' in this study. In contrast, benign tumors including Grade-I hemangiopericytoma and meningioma were not obvious in such behaviors. Moreover, when cultured in medium free of any anti-cancer drugs, cells from a recurrent Grade-III hemangiopericytoma that had been subjected to pre-operation adjuvant chemotherapy gradually shifted from non-mitotic proliferation to abnormal mitosis in the form of daughter number variation (DNV) and endomitosis, and eventually regular mitosis. Similarly, when treated with the anti-cancer drugs Epirubicin or Cisplatin, the cancer cell lines HeLa and A549 showed a shift from regular mitosis to abnormal mitosis, and further to non-mitosis as the dominant mode of proliferation with increasing drug concentrations. Upon removal of the drugs, the cells reversed back to regular mitosis with only minor occurrences of abnormal mitosis, accompanied by increased expression of the stem cell markers ALDH1, Sox, Oct4 and Nanog. CONCLUSIONS: The present study revealed that various types of malignant, but not benign, cancer cells exhibited cellular behaviors indicative of non-mitotic proliferation such as binary fission, which was typical of prokaryotic cell division, suggesting cell level atavism. Moreover, reversible transitions through the three modes of proliferation, i.e., mitosis, abnormal mitosis and non-mitosis, were observed when anticancer drug concentrations were grossly increased inducing non-mitosis or decreased favoring mitosis. Potential clinical significance of non-mitotic proliferation in cancer drug resistance and recurrence, and its relationship with cancer stem cells are worthy of further studies.

A gluten degrading probiotic Bacillus subtilis LZU-GM relieve adverse effect of gluten additive food and balances gut microbiota in mice.

Gluten accumulation damages the proximal small intestine and causes celiac disease (CeD) which has not been effectively treated except by using a gluten-free diet. In this study, strain Bacillus subtilis LZU-GM was isolated from Pakistani traditional fermented sourdough and could degrade 73.7% of gluten in 24 h in vitro. Strain LZU-GM was employed for practical application to investigate gluten degradation in mice models. The results showed that strain LZU-GM was colonized in mice and the survival rate was around 0.95 % (P < 0.0001). The gluten degradation was 3-fold higher in the small intestine of the strain LZU-GM treated mice group remaining 1511.96 ng/mL of gluten peptides than the untreated mice group (6500.38 ng/mL). Immunochemical analysis showed that gluten-treated mice established positive antigliadin antibodies (AGA) in serum (IgA, IgG, and anti-TG2 antibodies) as compared to the strain LZU-GM treatment group. Furthermore, the number of IFN-γ, TNF-α, IL-10, and COX-2 cells decrease in the lamina propria of the strain LZU-GM treatment group (P < 0.0001). Microbial community bar plot analysis showed that Lactobacillus, Dubosiella, and Enterococcus genera were restored and stabilized in the LZU-GM treatment group while Blautia and Ruminococcus were found lower. The oral gavage of probiotic strain LZU-GM might be useful for gluten metabolism in the intestine during digestion and would be a long-term dietary treatment for CeD management.

Lag Synchronization of Noisy and Nonnoisy Multiple Neurobiological Coupled FitzHugh-Nagumo Networks with and without Delayed Coupling.

This paper presents a methodology for synchronizing noisy and nonnoisy multiple coupled neurobiological FitzHugh-Nagumo (FHN) drive and slave neural networks with and without delayed coupling, under external electrical stimulation (EES), external disturbance, and variable parameters for each state of both FHN networks. Each network of neurons was configured by considering all aspects of real neurons communications in the brain, i.e., synapse and gap junctions. Novel adaptive control laws were developed and proposed that guarantee the synchronization of FHN neural networks in different configurations. The Lyapunov stability theory was utilized to analytically derive the sufficient conditions that ensure the synchronization of the FHN networks. The effectiveness and robustness of the proposed control laws were shown through different numerical simulations.

Analysis and quantification of naturally occurring aflatoxin B1 in dry fruits with subsequent physical and biological detoxification.

Current research involves extraction, identification and detoxification of mycotoxins from ten dry fruit samples. Mycotoxins were identified by high performance thin layer chromatography followed by physical and biological detoxification, analysed by HPLC. Three fungal species were observed after isolation including, Aspergillus niger, Aspergillus flavus and Fussarium sp. HP-TLC analysis revealed the presence of mycotoxin, aflatoxin B1 ranging from 0.000303-0.03636 mg/kg in all samples. Results were further analysed through various statistical tests. Detoxification methods proved to be cost effective and easily implementable. Concentration of aflatoxin B1 in pine nuts was reduced to 0.0043 mg/kg and 0.0039 mg/kg in dry dates through UV based detoxification. Solarisation reduced the concentration of aflatoxin B1 in figs to 0.0044 mg/kg. 90% aflatoxins were detoxified by UV treatment while Zingiber officinale powder detoxified 90% mycotoxin. This research concludes that the studied detoxification methods can be generalised on larger scale to benefit the dry fruit industry worldwide.

Mycotoxins in Zea mays, their quantification and HPLC analysis of physico-biological detoxification.

Present research delves in the isolation, extraction and identification of mycotoxins from ten corn samples collected from the northern province of Pakistan. Average concentration of aflatoxin B1 and B2 by HP-TLC found in all corn samples was 27.87 and 1.35 µg/kg, respectively. Following HP-TLC, detoxification of the identified and isolated mycotoxin was performed, which was analyzed by HPLC. Screening of mycoflora exhibited Aspergillus niger and Fusarium as the most dominant fungal strains. Aflatoxin B1 was physically detoxified under UV-Lamp and direct sunlight displaying detoxification percentage of 48% and 99%, respectively. Biological detoxification involved the use of botanicals such as neem leaves, garlic and ginger powder, which portrayed an approximate detoxification of 70% from corn samples. Current research concludes that the tested physical and biological methods can be easily adopted at field and storage rooms after the harvesting of crops to avoid fungal contamination and subsequent food spoilage.

Analysis of Fuel Alternative Products Obtained by the Pyrolysis of Diverse Types of Plastic Materials Isolated from a Dumpsite Origin in Pakistan.

The current energy crisis and waste management problems have compelled people to find alternatives to conventional non-renewable fuels and utilize waste to recover energy. Pyrolysis of plastics, which make up a considerable portion of municipal and industrial waste, has emerged as a feasible resolution to both satisfy our energy needs and mitigate the issue of plastic waste. This study was therefore conducted to find a solution for plastic waste management problems, as well as to find an alternative to mitigate the current energy crisis. Pyrolysis of five of the most commonly used plastics, polyethylene terephthalate (PET), high- and low-density polyethylene (HDPE, LDPE), polypropylene (PP), and polystyrene (PS), was executed in a pyrolytic reactor designed utilizing a cylindrical shaped stainless steel container with pressure and temperature gauges and a condenser to cool down the hydrocarbons produced. The liquid products collected were highly flammable and their chemical properties revealed them as fuel alternatives. Among them, the highest yield of fuel conversion (82%) was observed for HDPE followed by PP, PS, LDPE, PS, and PET (61.8%, 58.0%, 50.0%, and 11.0%, respectively). The calorific values of the products, 46.2, 46.2, 45.9, 42.8 and 42.4 MJ/kg for LPDE, PP, HPDE, PS, and PET, respectively, were comparable to those of diesel and gasoline. Spectroscopic and chromatographic analysis proved the presence of alkanes and alkenes with carbon number ranges of C9-C15, C9-C24, C10-C21, C10-C28, and C9-C17 for PP, PET, HDPE, LDPE, and PS, respectively. If implemented, the study will prove to be beneficial and contribute to mitigating the major energy and environmental issues of developing countries, as well as enhance entrepreneurship opportunities by replicating the process at small-scale and industrial levels.

Intrinsic and chemically-induced daughter number variations in cancer cell lines.

Multipolar mitosis was observed in cancer cells under mechanical stress or drug treatment. However, a comprehensive understanding of its basic properties and significance to cancer cell biology is lacking. In the present study, live-cell imaging was employed to investigate the division and nucleation patterns in four different cell lines. Multi-daughter divisions were observed in the three cancer cell lines HepG2, HeLa and A549, but not in the transformed non-cancer cell line RPE1. Multi-daughter mother cells displayed multi-nucleation, enlarged cell area, and prolonged division time. Under acidic pH or treatment with the anti-cancer drug 5-fluorouracil (5-FU) or the phytochemical compound wogonin, multi-daughter mitoses were increased to different extents in all three cancer cell lines, reaching as high as 16% of all mitoses. While less than 0.4% of the bi-daughter mitosis were followed by cell fusion events under the various treatment conditions, 50% or more of the multi-daughter mitoses were followed by fusion events at neutral, acidic or alkaline pH. These findings revealed a "Daughter Number Variation" (DNV) process in the cancer cells, with multi-daughter divisions in Stage 1 and cell fusions leading to the formation of cells containing up to five nuclei in Stage 2. The Stage 2-fusions were inhibited by 5-FU in A549 and HeLa, and by wogonin in A549, HeLa and HepG2. The parallel relationship between DNV frequency and malignancy among the different cell lines suggests that the inclusion of anti-fusion agents exemplified by wogonin and 5-FU could be beneficial in combinatory cancer chemotherapies.

Congruously designed eco-curative integrated farming model designing and employment for sustainable encompassments.

Eco-degradative features associated with the modern agriculture due to utilization of toxic agro-chemicals and intensified technologies need an urgent attention. Considering this need for eco-curativeness and eco-efficiency, current has for the first time employed an integrated farming system (IFS) through designing an appropriate assemblage of vegetables, poultry, and fish (VPF) and investigated its applied scale practicability in addition to the its role in the enhancement of the productivity and environmental quality maintenance. The practical employment of VPF model resulted in the remarkable improvement of soil fertility through an increment in the essential nutrient quantity. Physicochemical analysis of the soils expressed an improvement in the treated samples, i.e., pH (7.31), EC (0.92 dS/m), organic matter (2.97%), nitrogen (2.1 mg/kg), phosphorous (120.3 mg/kg), potassium (322 mg/kg), calcium (1482.0 mg/kg), and magnesium (471.5 mg/kg). Furthermore, ecological detoxification was expressed in form of lower heavy metals (HM) in the experimental soils. At the early plantation stage, HM concentration in the soils modified with nutrient-rich water signified considerably lower pattern with trend, i.e., Cd < Zn < Ni

Exploring perceptions of consanguineous unions with women from an East London community: analysis of discussion groups.

Consanguineous unions are relationships between blood relatives. This study explores the perceptions of consanguineous unions and risk of childhood disability and illness through the reported views and experiences of women in an ethnically diverse London community. This qualitative study utilised group discussions to elicit women's views and experiences. Field notes were recorded by independent note-takers in four group discussions. Field notes were coded manually and independently by two researchers who identified common themes for thematic analysis. Thirty-six women attended, of whom 20 identified as Asian Pakistani. Identified themes included variation in participants' views of consanguineous unions and associated health risks, the value of informed decisions and preferences for information distribution. Although participants had diverse opinions and experiences, they considered risk awareness to be vital for encouraging informed decisions in younger generations. This study highlights the importance of involving the community in efforts to increase awareness around consanguineous unions and genetic risk, emphasising the need for enabling educated choices and the value of co-developing educational efforts with the community.

A review on the elemental contents of Pakistani medicinal plants: Implications for folk medicines.

ETHNOPHARMACOLOGICAL RELEVANCE: Substantially, plants produce chemicals such as primary and secondary metabolites, which have significant applications in modern therapy. Indigenous people mostly rely on traditional medicines derived from medicinal plants. These plants have the capacity to absorb a variety of toxic elements. The ingestion of such plants for medicinal purpose can have imperative side effects. Hence, with regard to the toxicological consideration of medicinal plants, an effort has been made to review the elemental contents of ethno medicinally important plants of Pakistan and to highlight the existing gaps in knowledge of the safety and efficacy of traditional herbal medications. MATERIALS AND METHODS: Literature related to the elemental contents of ethno medicinal plants was acquired by utilizing electronic databases. We reviewed only macro-elemental and trace elemental contents of 69 medicinal plant taxa, which are traditionally used in Pakistan for the treatment of sundry ailments, including anemia, jaundice, cancer, piles, diarrhea, dysentery, headache, diabetes, asthma, blood purification, sedative and ulcer. RESULTS: A majority of plants showed elemental contents above the permissible levels as recommended by the World health organization (WHO). As an example, the concentrations of Cadmium (Cd) and Lead (Pb) were reportedly found higher than the WHO permissible levels in 43 and 42 medicinal plants, respectively. More specifically, the concentrations of Pb (54ppm: Silybum marianum) and Cd (5.25ppm: Artemisia herba-alba) were found highest in the Asteraceae family. CONCLUSIONS: The reported medicinal plants contain a higher amount of trace and toxic elements. Intake of these plants as traditional medicines may trigger the accumulation of trace and toxic elements in human bodies, which can cause different types of diseases. Thus, a clear understanding about the nature of toxic substances and factors affecting their concentrations in traditional medicines are essential prerequisites for efficacious herbal therapeutics with lesser or no side effects.

Northern hybridization: a proficient method for detection of small RNAs and microRNAs.

The significance of small RNAs, being an important part of the cellular machinery, is undeniable. Several techniques have been employed for detection of specific small RNAs. Among these techniques northern hybridization is the most popular due to its universal application to RNAs of various sizes. The general procedure involves separation of denatured RNA through gel electrophoresis and subsequent transfer and fixing to a membrane. RNA on the membrane is then detected using suitable labelled oligo-probes. Here we describe a method for detection of specific small RNAs, from an RNA pool, using sequence homology based oligonucleotide DNA or RNA probes.

Assessment of physicochemical parameters of wastewater samples.

Water samples from selected locations of Nullah Lai and Koh-e-Noor textile mill in the metropolitan city of Rawalpindi and Islamabad, Pakistan, were collected. Physicochemical parameters and heavy metals were determined using standard analytical procedures in comparison with sites, locations and subsequent interval of 3 months. The results of the physicochemical analysis at different locations of Nullah Lai and Koh-e-Noor textile mill with an interval of 3 months were obtained in the following range: pH (7.16-8.29), temperature (17.8-28.8 °C), conductivity (1,005-3,347 µS/m), TDS (754.3-2,519.5 mg/L), turbidity (272.8-487.05 NTU), total hardness (300-452 mg/L), nitrates (10.11-22.95 ppm), calcium (74.31-139.2 ppm), chloride (127.72-396.16 ppm), sulphate (15.97-87.38 ppm), NaCl (210.5-631.1 ppm), Ni (0.30-0.72 ppm), Cd (0.005-0.03 ppm), Cr (0.2-7.4 ppm), Pb (0.12-0.73 ppm), Zn (0.03-0.08 ppm) and Cu (0.01-0.06 ppm). The highest value of physicochemical parameters (compared with Nullah Lai) was obtained in locations of Koh-e-Noor textile mill. The results obtained exceeded the maximum allowable limit set by the World Health Organization for drinking purpose but can be used for irrigation purposes after suitable treatment and purification.

The hypersensitive response to tomato leaf curl New Delhi virus nuclear shuttle protein is inhibited by transcriptional activator protein.

The hypersensitive response (HR) is a common feature of plant disease resistance reactions and a type of programmed cell death (PCD). Many pathogens are able to modulate pathways involved in cell death. In contrast to animal viruses, inhibitors of PCD activity have not been identified for plant-infecting viruses. Previously, we have reported that the nuclear shuttle protein (NSP) of Tomato leaf curl New Delhi virus (ToLCNDV) induces an HR in Nicotiana tabacum and Lycopersicon esculentum plants when expressed under the control of the Cauliflower mosaic virus 35S promoter. However, HR is not evident in plants infected with ToLCNDV, suggesting that the virus encodes a factor (or factors) that counters this response. Analysis of all ToLCNDV-encoded genes pinpointed the transcriptional activator protein (TrAP) as the factor mediating the anti-HR effect. Deletion mutagenesis showed the central region of TrAP, containing a zinc finger domain and nuclear localization signal, to be important in inhibiting the HR. These results demonstrate that TrAP counters HR-induced cell death, the first such activity identified for a plant-infecting virus.

Analysis of variation in Alternaria alternata by pathogenicity and RAPD study.

Alternaria alternata were isolated and identified from root, foliage and soil of both wheat and rice crops and their aggressiveness was studied using aggressiveness analysis. Isolates of Alternaria alternata were genetically characterized using RAPD's. The investigations were based on surveys of wheat and rice crops in the rice-wheat cropping areas of Pakistan. The study showed that Alternaria alternata is root rot causing fungi and during root rot aggressiveness analysis the isolates showed higher aggressiveness on rice varieties than on wheat but in foliar aggressiveness the overall number of aggressive isolates was higher on wheat varieties than on rice. In genetic characterization Random Amplified Polymorphism DNA (RAPD) was used to study the polymorphism and genetic variation within the population of Alternaria alternata that established correlation between aggressiveness and genetical characters of fungi. Alternaria alternata tree is constructed based on the pattern of bands. This study highlighted the correlation between morphological, aggressiveness and genetic variations of Alternaria alternata.

The nuclear shuttle protein of Tomato leaf curl New Delhi virus is a pathogenicity determinant.

The role of the movement protein (MP) and nuclear shuttle protein (NSP) in the pathogenicity of Tomato leaf curl New Delhi virus (ToLCNDV), a bipartite begomovirus, was studied. Both genes were expressed in Nicotiana benthamiana, Nicotiana tabacum, and Lycopersicon esculentum plants with the Potato virus X (PVX) expression vector or by stable transformation of gene constructs under the control of the 35S promoter in N. tabacum. No phenotypic changes were observed in any of the three species when the MP was expressed from the PVX vector or constitutively expressed in transgenic plants. Expression of the ToLCNDV NSP from the PVX vector in N. benthamiana resulted in leaf curling that is typical of the disease symptoms caused by ToLCNDV in this species. Expression of NSP from PVX in N. tabacum and L. esculentum resulted in a hypersensitive response (HR), demonstrating that the ToLCVDV NSP is a target of host defense responses in these hosts. The NSP, when expressed as a transgene under the control of the 35S promoter, resulted in necrotic lesions in expanded leaves that initiated from a point and then spread across the leaf. The necrotic response was systemic in all the transgenic plants. Deletion of 100 amino acids from the C terminus did not compromise the HR response, suggesting that this region has no role in HR. Deletion of 60 or 100 amino acids from the N terminus of NSP abolished the HR response, suggesting that these sequences are required for the HR response. These findings demonstrate that the ToLCNDV NSP is a pathogenicity determinant as well as a target of host defense responses.