Calcium Crystals Research Articles

Additive coloration of crystals of calcium and cadmium fluorides

The particular features of additive coloration—formation of color centers as a result of annealing in a reducing atmosphere—have been considered for two crystals with fluorite structure, calcium and cadmium fluorides. The significant differences in the processes of coloration of these crystals are related to both the chemical reactions on their surface and the interaction (or absence of such) between the two components diffusing into the crystal bulk: anion vacancies and electrons.

Post-translational modifications regulate matrix Gla protein function: importance for inhibition of vascular smooth muscle cell calcification.

Matrix Gla protein (MGP) is a small vitamin K-dependent protein containing five gamma-carboxyglutamic acid (Gla) residues that are believed to be important in binding Ca(2+), calcium crystals and bone morphogenetic protein. In addition, MGP contains phosphorylated serine residues that may further regulate its activity. In vivo, MGP has been shown to be a potent inhibitor of vascular calcification; however, the precise molecular mechanism underlying the function of MGP is not yet fully understood. We investigated the effects of MGP in human vascular smooth muscle cell (VSMC) monolayers that undergo calcification after exposure to an increase in Ca(2+) concentration. Increased calcium salt deposition was found in cells treated with the vitamin K antagonist warfarin as compared to controls, whereas cells treated with vitamin K(1) showed decreased calcification as compared to controls. With conformation-specific antibodies, it was confirmed that warfarin treatment of VSMCs resulted in uncarboxylated (Gla-deficient) MGP. To specifically test the effects of MGP on VSMC calcification, we used full-length synthetic MGP and MGP-derived peptides representing various domains in MGP. Full length MGP, the gamma-carboxylated motif (Gla) (amino acids 35-54) and the phosphorylated serine motif (amino acids 3-15) inhibited calcification. Furthermore, we showed that the peptides were not taken up by VSMCs but bound to the cell surface and to vesicle-like structures. These data demonstrate that both gamma-glutamyl carboxylation and serine phosphorylation of MGP contribute to its function as a calcification inhibitor and that MGP may inhibit calcification via binding to VSMC-derived vesicles.

Cuff Tear Arthropathy

We investigated the role of two genes, ANKH and TNAP, in patients with cuff tear arthropathy. These genes encode proteins which regulate the extracellular concentration of inorganic pyrophosphate, fluctuations of which can lead to calcium crystal formation. Variants were detected by direct sequencing of DNA and their frequencies compared with healthy controls. The effect of variants on protein function was further studied by in vitro approaches. Variant genotypes were observed more frequently in the cases when compared with controls in ANKH (45% and 20%) and TNAP (32% and 9%). Variants in ANKH altered inorganic pyrophosphate (PPi) concentrations in transfected human chondrocytes. There was a higher mean serum concentration of TNAP detected in female patients compared with normal ranges. Cuff tear arthropathy is associated with variants in ANKH and TNAP that alter extracellular inorganic pyrophosphate concentrations causing calcium crystal deposition. This supports a theory that genetic variants predispose patients to primary crystal deposition which when combined with a massive rotator cuff tear leads to the development of arthritis.

Osterix Induces Osteogenic Gene Expression but not Differentiation in Primary Human Fetal Mesenchymal Stem Cells

The transcription factor osterix (Osx) is a key regulator of osteoblast differentiation and induces bone formation in embryonic but not adult stem cells. We investigated the effect of up-regulating Osx on an intermediate stem cell type, first trimester fetal mesenchymal stem cells (MSCs), which are more expandable than adult MSCs. Human fetal (hf ) MSCs were transduced with a lentiviral vector encoding human Osx. In undifferentiating MSCs cultures, forced expression of Osx stimulated osteopontin and alkaline phosphatase expression. However, Osx did not up-regulate osteocalcin, a late marker of osteoblast differentiation or result in extracellular calcium crystals, indicating that Osx does not directly mediate terminal differentiation in primary hfMSCs. To understand the downstream effects of Osx expression in primary hfMSCs, we next investigated the regulatory relationship between Osx, and the transcription factors Dlx5, Runx2, and Msx2. Osx induced Dlx5 but did not affect Runx2 and Msx2, whereas stealth ribonucleic acid interference of Osx inhibited Dlx5 without affecting expression of Runx2 and Msx2. In conclusion, Osx regulates osteogenic gene expression in hfMSCs but is insufficient to induce terminal osteogenic differentiation.

A case of massive kidney graft calcification that developed early in a child recipient with hyperparathyroidism

Abstract: A boy aged six yr and six months with end‐stage renal disease because of focal segmental glomerulosclerosis underwent cadaveric renal transplantation. Ischemic injury during the operation resulted in a severely dysfunctional graft. A renal graft biopsy was performed on post‐operative day (POD) 21. The biopsy specimen showed the presence of many intratubular calcium crystals, patchy tubular injury, and focal aggressive tubulointerstitial rejection. Pelvic computed tomography performed on POD 23 also showed prominent calcification in and around up to two‐thirds of the graft. After the biopsy, the graft function recovered, and hemodialysis was stopped on POD 22. A second graft biopsy was performed on POD 52. This biopsy specimen showed recovery from rejection, although the nephrocalcinosis remained. It was suggested that secondary hyperparathyroidism and hyperphosphatemia provoked nephrocalcinosis in this patient.

Fetuin-A, Valve Calcification, and Diabetes

In the current issue of Circulation , Ix et al demonstrate an inverse correlation between mitral and aortic valve calcification and serum fetuin-A levels in a cross-sectional study of 970 patients with coronary artery disease and without renal disease.1 Increased serum fetuin was significantly associated with diabetes mellitus, hypertriglyceridemia, serum albumin, and body mass index, with a weaker association with serum C-reactive protein, low-density lipoprotein cholesterol, and serum calcium. The link between aortic stenosis and fetuin held only in those without diabetes.1 The data highlight the complex interactions of fetuin with inflammation, insulin resistance, and tissue calcification. Furthermore, the results, though intriguing, underscore our lack of understanding of the cellular mechanisms of calcification in diverse pathological substrates, such as the degenerating aortic leaflet and mitral annulus. Article p 2533 Fetuin is a member of the cystatin superfamily of cysteine protease inhibitors, originally discovered as the major component of fetal bovine serum. It is a carrier for growth factors, binds to and inactivates transforming growth factor (TGF)–β and bone morphogenic protein, and is a major component of mineralized bone.2 Fetuin-A is also an acute-phase glycoprotein, produced in adults primarily in the liver, and is a powerful circulating inhibitor of hydroxyapatite formation. Mice that lack fetuin-A exhibit extensive soft-tissue calcification, which is accelerated on a mineral-rich diet, which suggests that fetuin-A acts to inhibit calcification systemically.3 In addition to its effects on mineralization, fetuin-A inhibits insulin receptor autophosphorylation and tyrosine kinase activity in vitro and in vivo. Fetuin-null knockout mice demonstrate improved insulin sensitivity and resistance to diet-induced obesity, and it has been postulated that the absence of fetuin in mice may contribute to the improvement of insulin sensitivity associated with aging.4 Clinically, serum fetuin-A is decreased in patients with moderate to severe chronic kidney disease, especially …

Update in calcium deposition diseases

Calcium pyrophosphate dihydrate and basic calcium phosphate crystals are common components of osteoarthritic synovial fluids and define subsets of patients with inflammatory or rapidly destructive arthritis. Recent literature concerning clinical and etiologic aspects of calcium pyrophosphate dihydrate and basic calcium phosphate crystal arthritis are reviewed. Recent literature reminds us of the propensity of calcium pyrophosphate dihydrate deposition disease to mimic other syndromes affecting the elderly. Several new studies reinforce the prevalence and significance of extra-articular calcium pyrophosphate dihydrate deposits, and demonstrate the presence of basic calcium phosphate-like whitlockite crystals in intervertebral discs. Current work serves to increase our appreciation for the complex role of the putative pyrophosphate transporter, ANKH, in healthy and diseased cartilage. The application of newer radiographic techniques to the diagnosis of calcium pyrophosphate dihydrate deposition disease holds promise for easier and more accurate identification of these crystal deposits in vivo. Work demonstrating the efficacy of a crystal poison in an animal model of osteoarthritis provides good evidence for a pathogenic role of calcium crystals in osteoarthritis, and hope for new therapies for these diseases. Continued work will further our understanding of these common crystals and their associated clinical syndromes.

Basic calcium phosphate crystals activate c- fos expression through a Ras/ERK dependent signaling mechanism

Diseases caused by calcium pyrophosphate dihydrate (CPPD) and basic calcium phosphate (BCP) crystals occur frequently in osteoarthritic joints. Both crystals induce mitogenesis, metalloproteinase synthesis and secretion by fibroblasts and chondrocytes, promoting degradation of articular tissue. We investigated the mechanism by which BCP activates the c- fos proto-oncogene, which has been shown to activate various matrix metalloproteinases ( MMPs). We demonstrate that BCP crystals induce c- fos expression primarily through a Ras/ERK-dependent signaling mechanism targeting two highly conserved regulatory binding sites, the serum response element (SRE) and the cAMP response element (CRE). These results establish a calcium crystal induced, calcium/calmodulin independent, signaling pathway in which BCP crystals activate Ras/MAPK, which can directly target an SRF-containing transcription factor complex, to induce fibroblasts to secrete metalloproteinases.

Tumoral Calcinosis of the Scalp: An Unusual Site for a Rare Tumor

Tumoral calcinosis is a rare calcifying disorder that is associated with deposition of calcium crystals in the periarticular tissues. The mass is most often around the hips, elbows, shoulders, and feet but may be occasionally found elsewhere. We report a case of multiple sporadic tumoral calcinoses in an adult male over the scalp. The scalp as a site of tumoral calcinosis has not been previously reported in adults. Previous surgical excisions done on two occasions had resulted in recurrence of the tumors. This report highlights the need to include tumoral calcinosis in the differential diagnosis of tumors of the scalp.

Metastatic pulmonary calcification in a dialysis patient: Case report and a review

A 19-year-old male presented with chest pain and dyspnea. He was anephric following nephrectomy for focal segmental glomerulosclerosis, had a subsequent failed transplant, and had been dialysis dependent for 3 years. Workup revealed hyperparathyroidism and an abnormal chest X-ray and computed tomography scan, significant for massive extra-skeletal pulmonary calcification. A markedly abnormal Technitium99 methylene diphosphonate (Tc99m-MDP) bone scan confirmed the clinical suspicion of metastatic pulmonary calcification. Metastatic pulmonary calcification (MPC) is common, occurring in 60% to 80% of dialysis patients on autopsy and bone scan series. It may lead to impaired oxygenation and restrictive lung disease. Typically, the calcium crystal is whitlockite rather than hydroxyapatite, which occurs in vascular calcification. Four major predisposing factors may contribute to MPC in dialysis patients. First, chronic acidosis leaches calcium from bone. Second, intermittent alkalosis favors deposition of calcium salts. Third, hyperparathyroidism tends to cause bone resorption and intracellular hypercalcemia. Finally, low glomerular filtration rate can cause hyperphosphatemia and an elevated calcium-phosphorus product. There may be other factors. Some authors suggest that the incidence of MPC in recent years may be lower due to improved dialysis techniques. The diagnosis is confirmed by biopsy, but can be suspected by typical findings on a Tc99m-MDP bone scan. Therapy is limited to ensuring adequate dialysis, correcting calcium-phosphorus product, and hyperparathyroidism; discontinuing vitamin D analogues may help. Conflicting reports show that transplantation may either improve or worsen the situation. MPC should be considered in dialysis patients who have characteristic abnormal chest radiography and/or pulmonary symptoms.

Molecular mechanisms mediating vascular calcification: Role of matrix Gla protein (Review Article)

Patients with chronic kidney disease (CKD) have a higher incidence of vascular calcification and a greatly increased risk of cardiovascular death. The mechanisms involved in the accelerated vascular calcification observed in CKD have recently become clearer, leading to the hypothesis that a lack of natural inhibitors of calcification may trigger calcium deposition. One of these inhibitory factors, matrix Gla protein (MGP), is the focus of the present review. MGP, originally isolated from bone, is a vitamin K-dependent protein that is also highly expressed by vascular smooth muscle cells. MGP has been confirmed as a calcification-inhibitor in numerous studies; however, its mechanism of action is not completely understood. It potentially acts in several ways to regulate calcium deposition including: (i) binding calcium ions and crystals; (ii) antagonizing bone morphogenetic protein and altering cell differentiation; (iii) binding to extracellular matrix components; and (iv) regulating apoptosis. Its expression is regulated by several factors including retinoic acid, vitamin D and extracellular calcium ions, and a reduced form of vitamin K (KH2) is important in maintaining MGP in an active form. Therefore, strategies aimed at increasing its expression and activity may be beneficial in tipping the balance in favour of inhibition of calcification in CKD.

Effect of some substances chemical on the crystallisation of calcium oxalate in vivo

Purpose: Epidemiological data collected during several decades that the majority of stones, up to 80% are composed of calcium oxalate (CaOx). This current study was aimed to look for an alternative treatment by using chemical substances on nephrolithiasic rabbits as a preventive against the development of urinary stone. The conditions of crystallisation and inhibition of oxalate calcium were studied at in vivo and in vitro. The experiment was conducted in normal and calcium oxalate (CaOx) nephrolithiasic abbits during three months. Several parameters were flowed every day including water intake, urinary volume and pH and crystalluria. Objectif: Our objective in this part of work was to determine the chemical inhibitors there act on the states of crystallization of calcium oxalate efficiently. A survey previous to the laboratory showed the in vitro inhibitory power of the chemical substances that we tested on the calcium oxalate. A comparison will be made between the two studies, only in the goal to confirm or to invalidate a possible effect of inhibition. The studied inhibitory substances are: the citric acid, Mg2*, Fe2*, F-. Methods: XVc divided the rabbits aged of 1 mois with a middleweight of 960g in two groups of 6. The second group was considered like witnesses. The two groups are nourished in industrial food. Result: The inhibition capacity of Fluoride, citrate, ferric and Magnesium ion were important on the aggregation and size of oxalate calcium crystals, but didn't inhibit completely this crystalline species.

Phosphocitrate, A Potential Therapeutic Agent for Calcium Crystal Deposition Diseases

The deposition of calcium-containing crystals in articular tissues is probably an under-recognized event. Clinical observations indicate that exaggerated and uniquely distributed cartilage degeneration is associated with these deposits. Perhaps the most compelling argument favoring a role for crystals in Osteoarthritis (OA) stems from their in vitro effects on articular tissues. Therapeutic options are limited and of compromised value for controlling and/or eliminating calcium crystal salt diseases. This review highlights past and present studies related to phosphocitrate (PC), a relatively unheralded compound with an ability to inhibit crystal nucleation, growth and aggregation of calcium salts, including basic calcium phosphate (a term including carbonate-substitute apatite, octacalcium phosphate, and tricalcium phosphate) and calcium pyrophosphate dihydrate. In addition, cell culture studies reveal that specific calcium phosphateinduced cellular events associated with osteoarthritis also are retarded by PC. Interest in the tetra-sodium PC form and the new Calcium/Sodium/PC (CaNaPC) salt has stemmed from their chemical characteristics and biological actions. In two instances, the CaNaPC has displayed superior inhibitory properties to that of the tetra-sodium salt. Using a calcergy animal model, a chemically-induced calcifying skin plaque in rats has been ameliorated while in cell culture studies, strong inhibition of calcium phosphate-DNA co-precipitates induced cell death has been noted. The assessed data indicate that either of the PC salts through their modes of action, could be useful as adjunct therapeutic treatment of crystal associated OA.

Morphological characteristics of Quercus variabilis charcoal prepared at different temperatures

The morphology of charcoals was investigated by scanning electron microscopy. Charcoal of Quercus variabilis was prepared in an electric furnace under nitrogen gas atmosphere at 400, 600, 800 and 1,000°C. Charring temperature greatly affects the structure of charcoal. In charcoal prepared at 400°C, most of the morphological characteristics remained relatively unchanged with the exception of the cell-wall layering. The cell walls appeared homogenous and glass-like. Above 400°C, there was an increase in cell-wall thinning and volumetric shrinkage with an increase in the charring temperature. These two factors were responsible for most of the observed changes in structure. Fracture surfaces became increasingly rough and disrupted. Vessel elements were increasingly distorted and tyloses disintegrated with increases in temperature. Parenchyma cells exhibited greater shrinkage except at the pits. This resulted in the appearance of distinctive small protuberances over the surface of the parenchyma cells. Rhomboidal calcium crystals were found to be abundant at all the temperatures studied, but at 800 and 1,000°C the crystals had a sponge-like appearance.

Disruption of clc-5 leads to a redistribution of annexin A2 and promotes calcium crystal agglomeration in collecting duct epithelial cells.

Mutations in CLCN5, which encodes the voltage-dependent Cl(-)/H(+)antiporter, CLC-5, cause Dent's disease. This disorder is characterized by low molecular-weight proteinuria, hypercalciuria, nephrocalcinosis and nephrolithiasis. Using a collecting duct cell model (mIMCD-3) in which endogenous clc-5 is disrupted by antisense clc-5 or overexpression of truncated clc-5, we demonstrate altered expression of the crystal adhesion molecule, annexin A2. Endogenously expressed annexin A2 is intracellular with limited plasma membrane localization. Following clc-5 disruption, there is both a marked increase in plasma membrane annexin A2 and an increase in cell surface crystal retention and agglomeration, which may be attenuated using pretreatment with anti-annexin A2 antibodies or wheat germ agglutinin lectin but not by concanavalin A. We hypothesize that in Dent's disease, endocytic failure leads to an accumulation at the plasma membrane of crystal-binding molecules that include annexin A2 leading to retention of calcium crystals and ultimately nephrocalcinosis and nephrolithiasis.

Tamm-Horsfall Protein is a Critical Renal Defense Factor Protecting Against Calcium Oxalate Crystal Formation

Tamm-Horsfall protein is a critical renal defense factor protecting against calcium oxalate crystal formation. Background The tubular fluid of the mammalian kidney is often supersaturated with mineral salts, but crystallization rarely occurs under normal conditions. The unique ability of the kidney to avoid harmful crystal formation has long been attributed to the inhibitory activity of the urinary macromolecules, although few in vivo studies have been carried out to examine this hypothesis. Here we examined the role of Tamm-Horsfall protein (THP), the principal urinary protein, in urinary defense against renal calcium crystal formation, using a THP knockout model that we recently developed. Methods Wild-type and THP knockout mice were examined for the spontaneous formation of renal calcium crystals using von Kossa staining. The susceptibility of these mice to experimentally induced renal crystal formation was evaluated by administering mice with ethylene glycol, a precursor of oxalate, and vitamin D 3 , which increases calcium absorption. Renal calcium crystals were visualized by von Kossa stain, dark field microscopy with polarized light and scanning electron microscopy. Results Inactivating the THP gene in mouse embryonic stem cells results in spontaneous formation of calcium crystals in adult kidneys. Excessive intake of calcium and oxalate, precursors of the most common type of human renal stones, dramatically increases both the frequency and the severity of renal calcium crystal formation in THP-deficient, but not in wild-type mice. Under high calcium/oxalate conditions, the absence of THP triggers a marked, adaptive induction in renal epithelial cells of osteopontin (OPN), a potent inhibitor of bone mineralization and vascular calcification. Thus, OPN may serve as an inducible inhibitor of calcium crystallization, whereas THP can serve as a constitutive and apparently more effective inhibitor. Conclusion These results provide the first in vivo evidence that THP is a critical urinary defense factor and suggest that its deficiency could be an important contributing factor in human nephrolithiasis, a condition afflicting tens of millions of people in the world annually.

Effects of OGP(10-14) and its derivative G38I on proliferation and differentiation of rat osteoblasts in vitro

To investigate the effects of the carboxyl end of osteogenic growth peptide (OGP)-OGP((10-14)) and its derivative G38I on the proliferation and differentiation of osteoblasts (OBs). Osteoblasts were isolated from the calvariae of newborn SD rats and cultured to G3. OGP((10-14)) or G38I of the concentrations of 10(-15) to 10(-7) mol/L were added to culture medium for 48 hours respectively. The number of cells was counted and MTT analysis was used to examine the proliferation of the cells. The ultrastructure of cells was investigated by electron microscopy. The osteoblasts of G3 were divided into experimental groups, treated with OGP((10-14)) or G38I of the concentration of 10(-11) mol/L for 48 hours, and control group. The alkaline phosphatase activity in the culture medium was measured. The protein expression level of type-I collagen was evaluated by immunohistochemistry. The core binding factor 1 (Cbfa1) and type-I collagen mRNA level of osteoblasts were determined by RT-PCR. With a biphasic effect on, OGP((10-14)) and G38I stimulated the number enhancement of OBs dose-dependently at low concentration and inhibited it at high concentration. The numbers of OB were the highest (37 x 10(4)/ml +/- 7 x 10(4)/ml and 30 x 10(4)/ml +/- 5 x 10(4)/ml respectively) when treated by OGP((10-14)) or G38I of the concentration of 10(-11)mol/L. The rough endoplasm net was flourishing and the secreting vesicle was abounding in the experimental cells. There was calcium crystal in the control cells. The activity of alkaline phosphatase in the culture medium of the OGP (10(-14)) and G38I groups were higher than that in the control group (4.47 U/g and 3.82 U/g vs 2.21 U/g). The protein expression level of type-I collagen was higher and the mRNA levels of Cbfa1 and type-I collagen were higher in the OGP((10-14)) and G38I groups were increased in the experimental groups in comparison with the control group (P < 0.05, P < 0.01, and P < 0.05). They stimulated cell number enhancement dose dependently at low concentration and followed by inhibition at high concentration. Just as the native OGP, OGP((10-14)) and its derivative G38I stimulate the proliferation of osteoblasts, and improve their activity, up-regulate the Cbfa1 and type-I collagen mRNA expression levels and increase the collagen synthesis, thus promoting the differentiation and osteogenic effect of osteoblasts.

Tumoral Calcinosis in Infants

Tumoral calcinosis is the deposition of calcium crystals and salts in the periarticular soft tissues. It commonly is seen as a complication of renal dialysis or in patients with a hereditary disposition. Occasionally, it is seen as an isolated condition with no known cause. Tumoral calcinosis usually affects adults and rarely, children. This disorder is extremely uncommon in infants. Only 13 cases have been reported. In this study three additional infants with tumoral calcinosis are reported, bringing the total to 16 cases. These 16 cases provide the opportunity to compare tumoral calcinosis in infants (younger than 18 months) with the disease in older patients. Although histologic and radiologic features in infants and adults are identical, some clinical features differ. In almost all infants, the disease develops without any predisposing factors, such as a family history. Surgical removal of lesions in infants is curative. By contrast, lesions in adults tend to recur after surgery.

(130) Dieffenbachia Calcium Oxalate Crystal Formation Affected by Cultivars, Nitrogen Rates, and Light Intensity

Tissue-culturedexplantsofDieffenbachiamaculate`Exotic Perfection', D.`Snow Flake', and D. × `Tropic Breeze' were grown on ebb-and-flow trays subirrigated with nitrogen (N) at 50, 200, or 800 mg·L-1 using a water-soluble fertilizer 17N–2.1P–15.7K for 10 weeks in a shaded greenhouse under a maximum photosynthetic photon flux density of 285 μmol·m-2·s-1. Plants were then transferred to interior rooms under a light level of 8 μmol·m-2·s-1. Samples of the midrib were taken from the first mature leaf of plants before being placed indoors and also from the first mature leaf of plants 8 months after growing indoors. Counts of calcium oxalate crystal idioblasts in cross-sections of the basal midrib using polarized light microscopy showed that the number of crystal idioblasts was higher in all three cultivars fertigated with 200 mg·L-1 N than those fertigated with either 50 or 800 mg·L-1 N. The number of crystal idioblasts in each cultivar grown under 8 μmol·m-2·s-1 was about 50% of the number detected when plants were grown under 285 μmol·m-2·s-1. `Snow Flake' had the highest number of crystal idioblasts with counts up to 60 per cross-section, whereas `Exotic Perfection' had the lowest with only 30 per cross-section. This study shows that in addition to cultivar differences, light intensity and N can significantly affect calcium crystal formation, and the highest number of crystal idioblasts occurred when Dieffenbachia cultivars were grown under optimum conditions.

CALCIUM OXALATE IN PLANTS: Formation and Function

Calcium oxalate (CaOx) crystals are distributed among all taxonomic levels of photosynthetic organisms from small algae to angiosperms and giant gymnosperms. Accumulation of crystals by these organisms can be substantial. Major functions of CaOx crystal formation in plants include high-capacity calcium (Ca) regulation and protection against herbivory. Ultrastructural and developmental analyses have demonstrated that this biomineralization process is not a simple random physical-chemical precipitation of endogenously synthesized oxalic acid and environmentally derived Ca. Instead, crystals are formed in specific shapes and sizes. Genetic regulation of CaOx formation is indicated by constancy of crystal morphology within species, cell specialization, and the remarkable coordination of crystal growth and cell expansion. Using a variety of approaches, researchers have begun to unravel the exquisite control mechanisms exerted by cells specialized for CaOx formation that include the machinery for uptake and accumulation of Ca, oxalic acid biosynthetic pathways, and regulation of crystal growth.