Autophagy promotes organelle clearance and organized cell separation of living root cap cells in Arabidopsis thaliana.

The root cap is a multilayered tissue covering the tip of a plant root that directs root growth through its unique functions, such as gravity sensing and rhizosphere interaction. To maintain the structure and function of the root cap, its constituent cells are constantly turned over through balanced cell division and cell detachment in the inner and outer cell layers, respectively. Upon displacement toward the outermost layer, columella cells at the central root cap domain functionally transition from gravity-sensing cells to secretory cells, but the mechanisms underlying this drastic cell fate transition are largely unknown. Here, using live-cell tracking microscopy, we show that organelles in the outermost cell layer undergo dramatic rearrangements. This rearrangement depends, at least partially, on spatiotemporally regulated activation of autophagy. Notably, this root cap autophagy does not lead to immediate cell death, but is instead necessary for organized separation of living root cap cells, highlighting a previously undescribed role of developmentally regulated autophagy in plants. This article has an associated 'The people behind the papers' interview.

Early sensing of phosphate deprivation triggers the formation of extra root cap cell layers via SOMBRERO through a process antagonized by auxin signaling.

KEY MESSAGE: The role of the root cap in the plant response to phosphate deprivation has been scarcely investigated. Here we describe early structural, physiological and molecular changes prior to the determinate growth program of the primary roots under low Pi and unveil a critical function of the transcription factor SOMBRERO in low Pi sensing. Mineral nutrient distribution in the soil is uneven and roots efficiently adapt to improve uptake and assimilation of sparingly available resources. Phosphate (Pi) accumulates in the upper layers and thus short and branched root systems proliferate to better exploit organic and inorganic Pi patches. Here we report an early adaptive response of the Arabidopsis primary root that precedes the entrance of the meristem into the determinate developmental program that is a hallmark of the low Pi sensing mechanism. In wild-type seedlings transferred to low Pi medium, the quiescent center domain in primary root tips increases as an early response, as revealed by WOX5:GFP expression and this correlates with a thicker root tip with extra root cap cell layers. The halted primary root growth in WT seedlings could be reversed upon transfer to medium supplemented with 250 µM Pi. Mutant and gene expression analysis indicates that auxin signaling negatively affects the cellular re-specification at the root tip and enabled identification of the transcription factor SOMBRERO as a critical element that orchestrates both the formation of extra root cap layers and primary root growth under Pi scarcity. Moreover, we provide evidence that low Pi-induced root thickening or the loss-of-function of SOMBRERO is associated with expression of phosphate transporters at the root tip. Our data uncover a developmental window where the root tip senses deprivation of a critical macronutrient to improve adaptation and surveillance.

Lateral Root Initiation and the Analysis of Gene Function Using Genome Editing with CRISPR in Arabidopsis.

Lateral root initiation is a post-embryonic process that requires the specification of a subset of pericycle cells adjacent to the xylem pole in the primary root into lateral root founder cells. The first visible event of lateral root initiation in Arabidopsis is the simultaneous migration of nuclei in neighbouring founder cells. Coinciding cell cycle activation is essential for founder cells in the pericycle to undergo formative divisions, resulting in the development of a lateral root primordium (LRP). The plant signalling molecule, auxin, is a major regulator of lateral root development; the understanding of the molecular mechanisms controlling lateral root initiation has progressed tremendously by the use of the Arabidopsis model and a continual improvement of molecular methodologies. Here, we provide an overview of the visible events, cell cycle regulators, and auxin signalling cascades related to the initiation of a new LRP. Furthermore, we highlight the potential of genome editing technology to analyse gene function in lateral root initiation, which provides an excellent model to answer fundamental developmental questions such as coordinated cell division, growth axis establishment as well as the specification of cell fate and cell polarity.

The growth of Arabidopsis primary root is repressed by several and diverse amino acids through auxin-dependent and independent mechanisms and MPK6 kinase activity.

Amino acids serve as structural monomers for protein synthesis and are considered important biostimulants for plants. In this report, the effects of all 20-L amino acids in Arabidopsis primary root growth were evaluated. 15 amino acids inhibited growth, being l-leucine (l-Leu), l-lysine (l-Lys), l-tryptophan (l-Trp), and l-glutamate (l-Glu) the most active, which repressed both cell division and elongation in primary roots. Comparisons of DR5:GFP expression and growth of WT Arabidopsis seedlings and several auxin response mutants including slr, axr1 and axr2 single mutants, arf7/arf19 double mutant and tir1/afb2/afb3 triple mutant, treated with inhibitory concentrations of l-Glu, l-Leu, l-Lys and l-Trp revealed gene-dependent, specific changes in auxin response. In addition, l- isomers of Glu, Leu and Lys, but not l-Trp diminished the GFP fluorescence of pPIN1::PIN1:GFP, pPIN2::PIN2:GFP, pPIN3::PIN3:GFP and pPIN7::PIN7:GFP constructs in root tips. MPK6 activity in roots was enhanced by amino acid treatment, being greater in response to l-Trp while mpk6 mutants supported cell division and elongation at high doses of l-Glu, l-Leu, l-Lys and l-Trp. We conclude that independently of their auxin modulating properties, amino acids signals converge in MPK6 to alter the Arabidopsis primary root growth.

Stem cell ageing of the root apical meristem of Arabidopsis thaliana.

Plants form new organs from pluripotent stem cells throughout their lives and under changing environmental conditions. In the Arabidopsis root meristem, a pool of stem cells surrounding a stem cell organizer, named Quiescent Center (QC), gives rise to the specific root tissues. Among them, the columella stem cell niche that gives rise to the gravity-sensing columella cells has been used as a model system to study stem cell regulation at the young seedling stage. However, little is known about the changes of the stem cell niche during later development. Here, we report that the columella stem cell niche undergoes pronounced histological and molecular reorganization as the plant progresses towards the adult stage. Commonly-used reporters for cellular states undergo re-patterning after an initial juvenile meristem phase. Furthermore, the responsiveness to the plant hormone abscisic acid, an integrator of stress response, strongly decreases. Many ageing effects are reminiscent of the loss-of-function phenotype of the central stem cell regulator WOX5 and can be explained by gradually decreasing WOX5 expression levels during ageing. Our results show that the architecture and central regulatory components of the root stem cell niche are already highly dynamic within the first weeks of development.

Calcium spikes, waves and oscillations in plant development and biotic interactions.

The calcium ion (Ca2+) is a universal signal in all eukaryotic cells. A fundamental question is how Ca2+, a simple cation, encodes complex information with high specificity. Extensive research has established a two-step process (encoding and decoding) that governs the specificity of Ca2+ signals. While the encoding mechanism entails a complex array of channels and transporters, the decoding process features a number of Ca2+ sensors and effectors that convert Ca2+ signals into cellular effects. Along this general paradigm, some signalling components may be highly conserved, but others are divergent among different organisms. In plant cells, Ca2+ participates in numerous signalling processes, and here we focus on the latest discoveries on Ca2+-encoding mechanisms in development and biotic interactions. In particular, we use examples such as polarized cell growth of pollen tube and root hair in which tip-focused Ca2+ oscillations specify the signalling events for rapid cell elongation. In plant-microbe interactions, Ca2+ spiking and oscillations hold the key to signalling specificity: while pathogens elicit cytoplasmic spiking, symbiotic microorganisms trigger nuclear Ca2+ oscillations. Herbivore attacks or mechanical wounding can trigger Ca2+ waves traveling a long distance to transmit and convert the local signal to a systemic defence program in the whole plant. What channels and transporters work together to carve out the spatial and temporal patterns of the Ca2+ fluctuations? This question has remained enigmatic for decades until recent studies uncovered Ca2+ channels that orchestrate specific Ca2+ signatures in each of these processes. Future work will further expand the toolkit for Ca2+-encoding mechanisms and place Ca2+ signalling steps into larger signalling networks.

Overexpression of three related root-cap outermost-cell-specific C2H2-type zinc-finger protein genes suppresses the growth of Arabidopsis in an EAR-motif-dependent manner.

The root meristem of Arabidopsis thaliana is protected by the root cap, the size of which is tightly regulated by the balance between the formative cell divisions and the dispersal of the outermost cells. We isolated an enhancer-tagged dominant mutant displaying the short and twisted root by the overexpression of ZINC-FINGER OF ARABIDOPSIS THALIANA1 (ZAT1) encoding an EAR motif-containing zinc-finger protein. The growth inhibition by ZAT1 was shared by ZAT4 and ZAT9, the ZAT1 homologues. The ZAT1 promoter was specifically active in the outermost cells of the root cap, in which ZAT1-GFP was localized when expressed by the ZAT1 promoter. The outermost cell-specific expression pattern of ZAT1 was not altered in the sombrero (smb) or smb bearskin1 (brn1) brn2 accumulating additional root-cap layers. In contrast, ZAT4-GFP and ZAT9- GFP fusion proteins were distributed to the inner root-cap cells in addition to the outermost cells where ZAT4 and ZAT9 promoters were active. Overexpression of ZAT1 induced the ectopic expression of PUTATIVE ASPARTIC PROTEASE3 involved in the programmed cell death. The EAR motif was essential for the growth inhibition by ZAT1. These results suggest that the three related ZATs might regulate the maturation of the outermost cells of the root cap. [BMB Reports 2020; 53(3): 160-165].

Activities of Chromatin Remodeling Factors and Histone Chaperones and Their Effects in Root Apical Meristem Development.

Eukaryotic genes are packaged into dynamic but stable chromatin structures to deal with transcriptional reprogramming and inheritance during development. Chromatin remodeling factors and histone chaperones are epigenetic factors that target nucleosomes and/or histones to establish and maintain proper chromatin structures during critical physiological processes such as DNA replication and transcriptional modulation. Root apical meristems are vital for plant root development. Regarding the well-characterized transcription factors involved in stem cell proliferation and differentiation, there is increasing evidence of the functional implications of epigenetic regulation in root apical meristem development. In this review, we focus on the activities of chromatin remodeling factors and histone chaperones in the root apical meristems of the model plant species Arabidopsis and rice.

Root cap size and shape influence responses to the physical strength of the growth medium in Arabidopsis thaliana primary roots.

During the progression of root in soil, root cap cells are the first to encounter obstacles and are known to sense environmental cues, thus making the root cap a potential mechanosensing site. In this study, a two-layered growth medium system was developed in order to study root responses to variations in the physical strength of the medium and the importance of the root cap in the establishment of these responses. Root growth and trajectory of primary roots of Arabidopsis seedlings were investigated using in vivo image analysis. After contact with the harder layer of the medium, the root either penetrated it or underwent rapid curvature, thus enabling reorientation of growth. We initially hypothesized that the root-cap structure would affect apex penetration and reorientation, with pointed caps facilitating and domed caps impeding root penetration. This hypothesis was investigated by analysing the responses of Arabidopsis mutants with altered root caps. The primary root of lines of the fez-2 mutant, which has fewer root-cap cell layers and a more pointed root cap than wild-type roots, showed impaired penetration ability. Conversely, smb-3 roots, which display a rectangular-shaped cap, showed enhanced penetration abilities. These results, which contradict our original hypothesis, reveal a role for resistance to buckling in determining root penetration abilities.

CRISPR-TSKO: A Technique for Efficient Mutagenesis in Specific Cell Types, Tissues, or Organs in Arabidopsis.

Detailed functional analyses of many fundamentally important plant genes via conventional loss-of-function approaches are impeded by the severe pleiotropic phenotypes resulting from these losses. In particular, mutations in genes that are required for basic cellular functions and/or reproduction often interfere with the generation of homozygous mutant plants, precluding further functional studies. To overcome this limitation, we devised a clustered regularly interspaced short palindromic repeats (CRISPR)-based tissue-specific knockout system, CRISPR-TSKO, enabling the generation of somatic mutations in particular plant cell types, tissues, and organs. In Arabidopsis (Arabidopsis thaliana), CRISPR-TSKO mutations in essential genes caused well-defined, localized phenotypes in the root cap, stomatal lineage, or entire lateral roots. The modular cloning system developed in this study allows for the efficient selection, identification, and functional analysis of mutant lines directly in the first transgenic generation. The efficacy of CRISPR-TSKO opens avenues for discovering and analyzing gene functions in the spatial and temporal contexts of plant life while avoiding the pleiotropic effects of system-wide losses of gene function.

Augmentation of root gravitropism by hypocotyl curvature in Brassica rapa seedlings.

Main Conclusion Root gravitropism of primary roots is assisted by curvature of the hypocotyl base. Root gravitropism is typically described as the sequence of signal perception, signal processing, and response that causes differential elongation and the establishment of a new gravitropic set-point angle. We describe two components of the graviresponse of Brassica seedlings that comprise a primary curvature of the root tip and a later onset but stronger curvature that occurs at the base of the hypocotyl. This second curvature is preceded by straightening of the tip region but leads to the completion of the alignment of the root axis. Curvature in both regions require a minimum displacement of 20 deg. The rate of tip curvature is a function of root length. After horizontal reorientation, tip curvature of five mm long roots curved twice as fast as 10 mm long roots (33.6 ±â€¯3.3 vs. 14.3 ±â€¯1.5 deg hr-1). The onset of curvature at the hypocotyl base is correlated with root length, but the rate of this curvature is independent of seedling length. Decapping of roots prevented tip curvature but the curvature at base of hypocotyl was unaffected. Endodermal cells at the root shoot junction show numerous, large and sedimenting amyloplasts, which likely serve as gravity sensors (statoliths). The amyloplasts at the hypocotyl were 3-4 µm in diameter, similar in size to those in the root cap, and twice the size of starch grains in the cortical layers of hypocotyl or elsewhere in the root. These data indicate that the root shoot reorientation of young seedlings is not limited to the root tip but includes more than one gravitropically responsive region.

Root extracellular traps versus neutrophil extracellular traps in host defence, a case of functional convergence?

The root cap releases cells that produce massive amounts of mucilage containing polysaccharides, proteoglycans, extracellular DNA (exDNA) and a variety of antimicrobial compounds. The released cells - known as border cells or border-like cells - and mucilage secretions form networks that are defined as root extracellular traps (RETs). RETs are important players in root immunity. In animals, phagocytes are some of the most abundant white blood cells in circulation and are very important for immunity. These cells combat pathogens through multiple defence mechanisms, including the release of exDNA-containing extracellular traps (ETs). Traps of neutrophil origin are abbreviated herein as NETs. Similar to phagocytes, plant root cap-originating cells actively contribute to frontline defence against pathogens. RETs and NETs are thus components of the plant and animal immune systems, respectively, that exhibit similar compositional and functional properties. Herein, we describe and discuss the formation, molecular composition and functional similarities of these similar but different extracellular traps.

Identification of Periplasmic Root-Cap Mucilage in Developing Columella Cells of Arabidopsis thaliana.

Plant roots secrete various substances with diverse functions against both plants and microbes in the rhizosphere. A major secretory substance is root-cap mucilage, whose functions have been well characterized, albeit mainly in crops. However, little is currently known about the developmental mechanisms of root-cap mucilage. Here, we show the accumulation and extrusion of root-cap mucilage in Arabidopsis. We found propidium iodide (PI) stainable structures between the plasma membrane and cell wall in the sixth layer of columella cells (c6) from the quiescent center. Ruthenium red staining and PI staining with calcium ions suggested that the structure comprises in part pectin polysaccharides. Electron microscopy revealed that the structure had a meshwork of electron-dense filaments that resembled periplasmic mucilage in other plants. In the c6 cells, we also observed many large vesicles with denser meshwork filaments to periplasmic mucilage, which likely mediate the transport of mucilage components. Extruded mucilage was observed outside a partially degraded cell wall in the c7 cells. Moreover, we found that the Class IIB NAC transcription factors BEARSKIN1 (BRN1) and BRN2, which are known to regulate the terminal differentiation of columella cells, were required for the efficient accumulation of root-cap mucilage in Arabidopsis. Taken together, our findings reveal the accumulation of and dynamic changes in periplasmic mucilage during columella cell development in Arabidopsis.

Xyloglucan and cellulose form molecular cross-bridges connecting root border cells in pea (Pisum sativum).

Pea (Pisum sativum) root cap releases a large number of living border cells that secrete abundant mucilage into the extracellular medium. Mucilage contains a complex mixture of polysaccharides, proteins and secondary metabolites important for its structure and function in defense. Unlike xyloglucan and cellulose, pectin and arabinogalactan proteins have been investigated in pea root and shown to be major components of border cell walls and mucilage. In this study, we investigated the occurrence of xyloglucan and cellulose in pea border cells and mucilage using cytochemical staining, immunocytochemistry and laser scanning confocal microscopy. Our data show that i) unlike cellulose, xyloglucan is highly present in the released mucilage as a dense fibrillary network enclosing border cells and ii) that xyloglucan and cellulose form molecular cross-bridges that tether cells and maintain them attached together. These findings suggest that secreted xyloglucan is essential for mucilage strengthening and border cell attachment and functioning.

The Root Cap Cuticle: A Cell Wall Structure for Seedling Establishment and Lateral Root Formation.

The root cap surrounding the tip of plant roots is thought to protect the delicate stem cells in the root meristem. We discovered that the first layer of root cap cells is covered by an electron-opaque cell wall modification resembling a plant cuticle. Cuticles are polyester-based protective structures considered exclusive to aerial plant organs. Mutations in cutin biosynthesis genes affect the composition and ultrastructure of this cuticular structure, confirming its cutin-like characteristics. Strikingly, targeted degradation of the root cap cuticle causes a hypersensitivity to abiotic stresses during seedling establishment. Furthermore, lateral root primordia also display a cuticle that, when defective, causes delayed outgrowth and organ deformations, suggesting that it facilitates lateral root emergence. Our results show that the previously unrecognized root cap cuticle protects the root meristem during the critical phase of seedling establishment and promotes the efficient formation of lateral roots.

Cytogenetics and morphological delimitation between three species of Passiflora L. (subgenus Distephana Cervi).

Several studies on cytogenetic characterisation of passion flowers are helpful to elucidate doubts about taxa relationships, delimitation and classification into more coherent groups based on karyomorphological data. Molecular and conventional cytogenetic techniques were applied to three Passiflora species with red flowers, P. coccinea, P. vitifolia and P. tholozanii, for species karyotype relationships. Additionally, for descriptive morphology, were used flowers, leaves and seeds. Results describe for the first time the karyomorphological and chromosome number (2n = 18) for P. tholozanii. anova was performed (P < 0.05) and statistical significance for average chromosome size (CV: 16.53%) between species. Genomic in situ hybridisation (GISH) proved relationships between P. coccinea and P. tholozanii, which suggests a common origin, however, we could not identify hybridisation between genomic probes from P. vitifolia in P. tholozanii chromosomes. Among the species analysed, P. tholozanii has great similarity in karyotypic and morphology to P. coccinea but not to P. vitifolia. We suggest the inclusion of P. tholozanii in the same subgenus and section as P. coccinea based on the similarity in karyomorphological and morphological traits between the species. Additionally, GISH might indicate a common or hybrid origin of P. tholozanii.

Depicting the physiological and ultrastructural responses of soybean plants to Al stress conditions.

Aluminium (Al) is a toxic element for plants living in soils with acidic pH values, and it causes reductions in the roots and shoots development. High Al concentrations can cause physiological and structural changes, leading to symptoms of toxicity in plant tissue. The aim of this study was to describe the Al toxicity in soybean plants through physiological, nutritional, and ultrastructure analyses. Plants were grown in nutrient solution containing increasing Al concentrations (0; 0.05; 0.1; 1.0, 2.0 and 4.0 mmol L-1). The Al toxicity in the soybean plants was characterized by nutritional, anatomical, physiological, and biochemical analyses. The carbon dioxide assimilation rates and stomatal conductance were not affected by the Al. However, the capacity for internal carbon use decreased, and the transpiration rate increased, resulting in increased root biomass at the lowest Al concentration in the nutrient solution. The soybean plants exposed to the highest Al concentration exhibited lower root and shoot biomass. The nitrate reductase and urease activities decreased with the increasing Al concentration, indicating that nitrogen metabolism was halted. The superoxide dismutase and peroxidase activities increased with the increasing Al availability in the nutrient solution, and they were higher in the roots, showing their role in Al detoxification. Despite presenting external lesions characterized by a damaged root cap, the root xylem and phloem diameters were not affected by the Al. However, the leaf xylem diameter showed ultrastructural alterations under higher Al concentrations in nutrient solution. These results have contributed to our understanding of several physiological, biochemical and histological mechanisms of Al toxicity in soybean plants.

Aluminium-induced cell death requires upregulation of NtVPE1 gene coding vacuolar processing enzyme in tobacco (Nicotiana tabacum L.).

Cell death mechanism triggered by aluminium (Al) ion was investigated at root apex of tobacco (cultivar Bright Yellow) and in cultured tobacco cell line BY-2 derived from Bright Yellow, focusing on VPE genes (NtVPE1a, NtVPE1b, NtVPE2, NtVPE3). Cell death was detected as a loss of integrity of the plasma membrane by vital staining with fluorescein diacetate (in root apex) and Evans blue (in BY-2), respectively. At root apex, the upregulation of gene expression of VPE1a and VPE1b was observed significantly after 9h of Al exposure in parallel with an enhancement of cell death, while the upregulation of VPE2 and VPE3 were observed later. Similarly, in BY-2 cells, the upregulation of VPE1a and VPE1b and the enhancement of cell death were synchronously observed after 3-h exposure to Al, while the upregulation of VPE2 and VPE3 occurred later. RNA interference (RNAi) lines of each of the VPEs were constructed in BY-2 cells. Comparative studies between wild-type and the RNAi lines indicated that both Al-enhanced VPE activity and Al-induced cell death were significantly suppressed in the RNAi lines of VPE1 (dual suppressor of VPE1a and VPE1b), but not in the RNAi lines of VPE2 and that of VPE3. Taken together, we conclude that the upregulation of VPE1 gene expression and following enhancement of VPE activity under Al stress cause cell death in actively growing or elongating cells of tobacco.

Effects of Bacillus subtilis on some physiological and biochemical parameters of Triticum aestivum L. (wheat) under salinity.

Endophytic strain Bacillus subtilis (B. subtilis) 10-4, producing indole-3-acetic acid (IAA) and siderofores but not active in phosphate solubilization, exerted a protective effect on Triticum aestivum L. (wheat) plant grown under salinity (2% NaCl) stress. Exposure to salt stress resulted in an essential increase of proline (Pro) and malondialdehyde (MDA) level in the seedlings. At the same time the seedlings inoculated with B. subtilis 10-4 were characterized by decreased level of stress-induced Pro and MDA accumulation. It was revealed that both B. subtilis 10-4 and salinity caused increase in the content of endogenous salicylic acid (SA) in wheat seedlings as compared to SA content in the control, while B. subtilis 10-4 suppressed stress-induced SA accumulation. Water storage capacity (WSC) in leaf tissues was increased and stress-induced hydrolysis of statolite starch in root cap cells of the germinal roots was reduced by B. subtilis 10-4. The obtained data indicated that the activation of the defense reactions induced by B. subtilis 10-4 induced defense reactions may be connected with their ability to decrease the level of stress-induced oxidative and osmotic stress in seedlings and with the increase of endogenous SA level that can make a significant contribution to the implementation of the protective effect of B. subtilis 10-4 and is manifested in the improvement of plant growth, WSC of leaves and slowing down of the process of statolite starch hydrolysis under salinity.

Haematopoietic effects of Angelica sinensis root cap polysaccharides against lisinopril-induced anaemia in albino rats.

CONTEXT: Angelica sinensis L. (Umbelliferae) has medicinal properties. OBJECTIVES: The present study evaluates the haematopoietic effects of A. sinensis polysaccharides (ASP) against lisinopril-induced anaemia. MATERIALS AND METHODS: Thirty healthy adult male albino rats were randomly divided into five groups (n = 6). Group I was control group. Group II was treated with angiotensin-converting enzyme inhibitor (ACEI, 20 mg/kg/day) to induce anaemia. In group III, erythropoietin (EPO, 100 IU/kg/each) was administered in combination with ACEI. Group IV was treated with ASP (1 g/kg/day), extracted from A. sinensis root caps. In Group V, ASP (1 g/kg/day) was treated with ACEI. After 28 days, blood and tissue samples were collected for haematological and histopathological analysis, respectively. RESULTS: The results showed that ACEI significantly reduced the haemoglobin (Hb, 10.0 g/dL), packed cell volume (PCV, 39.5%), red blood cells (RBCs, 6.2 million/mm3), mean corpuscular volume (MCV, 53.5 fL) and mean corpuscular haemoglobin (MCH, 16.2 pg/cell) values. In the group treated with ASP, the Hb (13.7 g/dL) and RBCs (7.8 million/mm3) increased significantly (p < 0.05). The combination of ASP and ACEI led to the significant (p < 0.05) reduction in Hb (10.7 g/dL), PCV (33.3%), RBCs (6.0 million/mm3), MCV (54.42 fL) and MCH (16.44 pg/cell) values. While histopathological examination of the liver and kidney cells showed a mild degree of toxicity in the ASP-treated group. CONCLUSION: ASP has a potentiating effect on haematological parameters when given alone. However, when administered simultaneously with lisinopril, it showed an unfavourable effect with more complicated anaemia so it should not be used with ACEIs.