BroadfordingScience

^{Articles > The Difference between Planarian’s Regeneration in Three Types of Water}

_{Date Published: 1 November 2024}

The Difference between Planarian’s Regeneration in Three Types of Water

Geon Choi, Hyunjun Shim, and Younghyun Lee

---

Abstract

This research utilized three types of water, Spring Water, Lake Water, and Deionized Water, to quantitatively measure the regeneration of planarians in each water sample. Spring and Deionized water were collected from the GVCS Broadfording cafeteria and chemistry laboratory. The media storage bottles were autoclaved under pressure to sterilize before containing water samples. The experiment’s initial hypothesis was that the spring water regeneration would work best, and stream water would deter the regeneration. Unlike the hypothesis, the result indicated that the planarian’s regeneration worked best in the stream water; the deionized water was least suitable for the regeneration of the planarian.

Introduction:

Cell proliferation and differentiation regulation in adult tissues is important to many fundamental biological processes, including wound repair, regeneration, and cancer treatment. Tumor formation has been proposed as a pathological form of tissue regeneration. Both regeneration and tumor formation involve activating stem cells, which proliferate to replace lost tissues in response to injury or form tumors in a pathological context. Therefore, understanding the molecular mechanisms governing tissue regeneration can provide valuable insights into the dysregulation of cell proliferation in adult tissues.

Planarians are a well-characterized animal regeneration model and are accessible to the scientific literature. Planarians demonstrate remarkable regenerative abilities. Planarians show an amazing ability to regenerate into a new, complete organism. This impressive regenerative capacity is due to the following reasons: Large numbers of adult pluripotent stem cells, called neuroblasts, are carried in their bodies, which is why they can regenerate. For example, when a wound is inflicted, such as an amputation, the neoplastic cells are activated and immediately proliferate at the wound site, forming a specialized tissue called blastema. 

Planarians may regenerate their bodies from small tissue fragments to the entire body. This regeneration makes a planarian special organism. The capabilities of regeneration have five big steps:

First, for Wound Closure, planarians immediately initiate wound closure after injury by contracting muscles around the wound site. This process is crucial for preventing excessive bleeding and protecting the internal environment from external threats.

Second, for the Formation of Blastem, a group of undifferentiated cells known as the blastema forms at the injury site. These cells possess the remarkable capability to differentiate into various cell types needed for the regeneration process.

Third, cell proliferation and differentiation occur rapidly. Stem cells in the planarian’s body divide and specialize, generating the diverse cell types necessary to reconstruct lost or damaged body parts.

Fourth, as cell proliferation and differentiation progress, signaling molecules such as Wnt, BMP, and FGF play crucial roles in orchestrating the spatial organization and patterning of newly formed tissues. These molecules help establish the correct anatomical axes, including the anterior-posterior and dorsal-ventral axes, ensuring proper body plan restoration.

Finally, the newly formed tissues integrate seamlessly with existing structures, leading to the maturation of regenerated body parts. This phase involves the refinement of tissue structures, functional restoration, and establishing connectivity with the nervous system to ensure full functionality of the regenerated body parts.

The pH level of stream water, influenced by factors such as geological composition, surrounding vegetation, and anthropogenic inputs, serves as a key determinant of chemical equilibrium and biological activity. Optimal pH levels within the physiological range are essential for maintaining cellular homeostasis and enzymatic function in aquatic organisms. Deviations from this range can disrupt cellular processes, including those involved in tissue repair and regeneration, by altering protein structure, enzyme activity, and ion balance.

Stream water’s oxygenation status profoundly impacts aquatic organisms’ cellular metabolism and energy production. Dissolved oxygen levels, influenced by temperature, water flow, and biological activity, directly affect aerobic respiration, the primary mode of energy generation in most organisms. Adequate oxygenation facilitates efficient ATP production, supporting cellular functions essential for tissue repair and regeneration. Conversely, hypoxic or anoxic conditions can impede cellular metabolism, leading to metabolic stress and compromising regenerative capacities.

Stream water is composed of various chemicals from natural and anthropogenic sources. The most common chemicals in water pollutants include heavy metals such as mercury and arsenic, which can originate from mining activities, agricultural runoff, atmospheric deposition, and waste disposal. 

Arsenic is a naturally occurring metalloid found in the earth’s crust. It exists in two forms: inorganic and organic. Arsenic becomes more toxic when it exists in an inorganic form. This chemical enters water through runoff (mining operations), atmospheric deposition, and improper waste disposal. 

Mercury is a heavy metal in various forms: elemental mercury, inorganic compounds, and methylmercury. This chemical may intrude through mining, industrial processes, and the atmosphere. 

Deionized water (DI water) removes most mineral ions, such as sodium, calcium, and iron. This water is used in laboratories, medical procedures, and industrial processes. Sea animals require specific amounts of ions, which can be found in seawater; however, due to the lack of minerals and ions that sea animals need, they cannot live in DI water. 

Materials:

Deionized  Water		From the school lab’s Deionized water filter system.  The DI process leverages specially manufactured ion-exchange resins that exchange hydrogen (H+) and hydroxyl (OH-) ions for dissolved minerals and then recombine to form water (H2O).
Spring Water		Deer Park® Brand Natural Spring Water 16.9 oz
Stream Water		The water was collected near Hagerstown’s Conococheague Creek.  Filtering Apparatus includes:Filtering funnelOne-hole stopperGlass fritFiltering flask Microdisk Membrane Filters that were sterilized were used.
Autoclave		All the bottles were sterilized by autoclave. The items were heated to 120 °C for 60 minutes to kill bacteria in the bottles.
PYREX Autoclavable Bottles (250 mL)		Vented membrane cap with 0.22 µm PTFE hydrophobic membrane.Red PBT (polybutylene terephthalate) solid caps and ETFE pouring rings for dry heat sterilization (180°C).PBT caps with silicone septa are used for syringe sampling or the introduction of reagents.

Experimental Process: 

First Trial

The only conditions that allowed the planarians to live in the first place were in dark spaces and to change the water periodically. Before remodeling the planarians, there was an accident in culturing the planarians twice. First, tap water did not meet the appropriate conditions for the systems’ living conditions, culturing the planarians. 

Second Trial

In the second attempt, we tried to clean and cultivate the living spaces using only spring water, but within a week, we saw all planarians dead.

Third Trial 

In the third experiment, planarians were divided into three groups and raised in different types of water: distilled (DI), spring, and lake. At the start of the full-scale experiment, all tools were sterilized and cleaned with DI water. Water was changed thrice a week, and freshwater was collected directly from the stream weekly for the group in valley water. No planarians died during this period. Some planarians from each bottle were transferred to Petri dishes and cut into two or more pieces, with both the head and torso showing movement. A dark environment was prepared for the planarians, and water was changed every two days after washing. During the third week, the planarians were fed crushed egg yolks and transferred to new bottles 30 minutes later. After feeding, the planarians in DI water started to die gradually. Due to difficulties in collecting valley water, the experiment was concluded. The properties and regeneration processes of the planarians in each type of water were then analyzed and compared with the initial hypothesis.

Results:

Table 1: Planarian Generation by each type of water

	Deionized Water	Spring water	Stream Water
4.12 ~ 4.19
4.19 ~ 4.26
4.26 ~ 5.3

Table 1 shows the number of planarian regeneration processes in each type of water: deionized water, spring water, and stream water. Pictures of data were collected every seven days beginning on April twelve. Two individual planarians were cultured from each system group separately in beakers. The planarians were divided into two halves and divided into observations, bodies, and trunks. Three weeks after cutting planarians, in the beaker of deionized water, all planarians were found dead. On the other hand, each planarian was found alive in the beaker of spring water and stream water. For the difference between planarians in spring and stream water, planarians in the stream water seemed to regenerate more than in the spring water. 

The experiment’s results reveal significant disparities in planarian survival and regeneration capabilities across different water environments. In deionized (DI) water, all planarians were found deceased three weeks after the experiment’s initiation, indicating a complete lack of survivability in this environment. Conversely, planarians cultured in spring and stream water survived the same observation period.

Discussions:

The observed differences in survival rates can be attributed to the chemical composition and environmental conditions of the respective water sources. It is hypothesized that the deionized water used in the experiment contained the least amount of contaminant and lacked essential nutrients necessary for planarian survival and regeneration.

The capacity of planarian regeneration observed in stream water compared to spring water signifies that the environmental conditions and chemical composition of stream water from Conococheague Creek in Hagerstown, MD, may provide specific advantages for tissue repair and regeneration processes. In addition, the dynamic flow and circulation patterns within streams facilitate the removal of metabolic wastes and provide better oxygenation, creating optimal conditions for cellular metabolism and regeneration.

pH levels and dissolved oxygen concentrations may differ between stream water and spring water, with stream water potentially offering more stable and favorable conditions for planarian regeneration. The continuous flow of water in streams may help maintain stable environmental parameters. In contrast, spring water may lack the dynamic environmental fluctuations necessary to stimulate robust regenerative responses in planarians.

Conclusion:

The research was conducted to investigate the difference in planarian regeneration in three types of water. As introduced earlier, planarians demonstrate remarkable regenerative abilities. To eliminate the possible contamination error in the planaria cultivation, the laboratory autoclave was used at a sterilization temperature to kill bacteria in the bottles. Samples of spring, deionized, stream water were taken from GVCS broadfording cafeteria, laboratory, and Conococheague Creek. It is assumed that regeneration in spring water would work the best and the least in stream water.

The result was unexpected; for the difference between regeneration of planarians in spring and stream water, planarians in the stream water were observed to regenerate more than in the spring water. In the beaker of deionized water, all planarians were found dead. The data rejects the initial hypothesis by stream water’s abundant oxygenation, nutrients, and optimal conditions.

This experiment demonstrates that water quality significantly influences planarian regeneration, with stream water yielding superior outcomes to spring water. The detrimental effects observed in deionized water highlight the importance of considering water composition in research settings and aquatic habitats.